Publications by authors named "Xingming Shi"

40 Publications

The Glucocorticoid Receptor in Osterix-Expressing Cells Regulates Bone Mass, Bone Marrow Adipose Tissue, and Systemic Metabolism in Female Mice During Aging.

J Bone Miner Res 2021 Nov 8. Epub 2021 Nov 8.

Department of Cellular Biology and Anatomy, Augusta University, Augusta, GA, USA.

Hallmarks of aging-associated osteoporosis include bone loss, bone marrow adipose tissue (BMAT) expansion, and impaired osteoblast function. Endogenous glucocorticoid levels increase with age, and elevated glucocorticoid signaling, associated with chronic stress and dysregulated metabolism, can have a deleterious effect on bone mass. Canonical glucocorticoid signaling through the glucocorticoid receptor (GR) was recently investigated as a mediator of osteoporosis during the stress of chronic caloric restriction. To address the role of the GR in an aging-associated osteoporotic phenotype, the current study utilized female GR conditional knockout (GR-CKO; GR :Osx-Cre+) mice and control littermates on the C57BL/6 background aged to 21 months and studied in comparison to young (3- and 6-month-old) mice. GR deficiency in Osx-expressing cells led to low bone mass and BMAT accumulation that persisted with aging. Surprisingly, however, GR-CKO mice also exhibited alterations in muscle mass (reduced % lean mass and soleus fiber size), accompanied by reduced voluntary physical activity, and also exhibited higher whole-body metabolic rate and elevated blood pressure. Moreover, increased lipid storage was observed in GR-CKO osteoblastic cultures in a glucocorticoid-dependent fashion despite genetic deletion of the GR, and could be reversed via pharmacological inhibition of the mineralocorticoid receptor (MR). These findings provide evidence of a role for the GR (and possibly the MR) in facilitating healthy bone maintenance with aging in females. The effects of GR-deficient bone on whole-body physiology also demonstrate the importance of bone as an endocrine organ and suggest evidence for compensatory mechanisms that facilitate glucocorticoid signaling in the absence of osteoblastic GR function; these represent new avenues of research that may improve understanding of glucocorticoid signaling in bone toward the development of novel osteogenic agents. © 2021 American Society for Bone and Mineral Research (ASBMR).
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http://dx.doi.org/10.1002/jbmr.4468DOI Listing
November 2021

Age-associated changes in microRNAs affect the differentiation potential of human mesenchymal stem cells: Novel role of miR-29b-1-5p expression.

Bone 2021 12 14;153:116154. Epub 2021 Aug 14.

Department of Pathology and Laboratory Medicine, Medical University of South Carolina, Charleston, SC 29403, United States of America; Ralph H. Johnson Veterans Affairs Medical Center, Charleston, SC 29403, United States of America; Department of Cellular Biology and Anatomy, Medical College of Georgia, Augusta University, Augusta, GA 30912, United States of America; Center for Healthy Aging, Medical College of Georgia, Augusta University, Augusta, GA 30912, United States of America. Electronic address:

Age-associated osteoporosis is widely accepted as involving the disruption of osteogenic stem cell populations and their functioning. Maintenance of the local bone marrow (BM) microenvironment is critical for regulating proliferation and differentiation of the multipotent BM mesenchymal stromal/stem cell (BMSC) population with age. The potential role of microRNAs (miRNAs) in modulating BMSCs and the BM microenvironment has recently gained attention. However, miRNAs expressed in rapidly isolated BMSCs that are naïve to the non-physiologic standard tissue culture conditions and reflect a more accurate in vivo profile have not yet been reported. Here we directly isolated CD271 positive (+) BMSCs within hours from human surgical BM aspirates without culturing and performed microarray analysis to identify the age-associated changes in BMSC miRNA expression. One hundred and two miRNAs showed differential expression with aging. Target prediction and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway analyses revealed that the up-regulated miRNAs targeting genes in bone development pathways were considerably enriched. Among the differentially up-regulated miRNAs the novel passenger strand miR-29b-1-5p was abundantly expressed as a mature functional miRNA with aging. This suggests a critical arm-switching mechanism regulates the expression of the miR-29b-1-5p/3p pair shifting the normally degraded arm, miR-29b-1-5p, to be the dominantly expressed miRNA of the pair in aging. The normal guide strand miR-29b-1-3p is known to act as a pro-osteogenic miRNA. On the other hand, overexpression of the passenger strand miR-29b-1-5p in culture-expanded CD271+ BMSCs significantly down-regulated the expression of stromal cell-derived factor 1 (CXCL12)/ C-X-C chemokine receptor type 4 (SDF-1(CXCL12)/CXCR4) axis and other osteogenic genes including bone morphogenetic protein-2 (BMP-2) and runt-related transcription factor 2 (RUNX2). In contrast, blocking of miR-29b-1-5p function using an antagomir inhibitor up-regulated expression of BMP-2 and RUNX2 genes. Functional assays confirmed that miR-29b-1-5p negatively regulates BMSC osteogenesis in vitro. These novel findings provide evidence of a pathogenic anti-osteogenic role for miR-29b-1-5p and other miRNAs in age-related defects in osteogenesis and bone regeneration.
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http://dx.doi.org/10.1016/j.bone.2021.116154DOI Listing
December 2021

Photobiomodulation has rejuvenating effects on aged bone marrow mesenchymal stem cells.

Sci Rep 2021 06 22;11(1):13067. Epub 2021 Jun 22.

Department of Neuroscience and Regenerative Medicine, Medical College of Georgia, Augusta University, 1120 15th Street, CA-2004, Augusta, GA, 30912, USA.

The plasticity and proliferative capacity of stem cells decrease with aging, compromising their tissue regenerative potential and therapeutic applications. This decline is directly linked to mitochondrial dysfunction. Here, we present an effective strategy to reverse aging of mouse bone marrow mesenchymal stem cells (BM-MSCs) by restoring their mitochondrial functionality using photobiomodulation (PBM) therapy. Following the characterization of young and aged MSCs, our results show that a near-infrared PBM treatment delivering 3 J/cm is the most effective modality for improving mitochondrial functionality and aging markers. Furthermore, our results unveil that young and aged MSCs respond differently to the same modality of PBM: whereas the beneficial effect of a single PBM treatment dissipates within 7 h in aged stem cells, it is lasting in young ones. Nevertheless, by applying three consecutive treatments at 24-h intervals, we were able to obtain a lasting rejuvenating effect on aged MSCs. Our findings are of particular significance for improving autologous stem cell transplantation in older individuals who need such therapies most.
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http://dx.doi.org/10.1038/s41598-021-92584-3DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8219765PMC
June 2021

Ameliorative Effects of Component Chinese Medicine From and , a Traditional Herb Pair, on Uterine Leiomyoma in a Rat Model.

Front Public Health 2021 28;9:674357. Epub 2021 May 28.

Basic Medicine College, Chengdu University of Traditional Chinese Medicine, Chengdu, China.

Uterine leiomyoma (UL), common benign tumors in women of child-bearing age, are believed to be caused mainly by Qi stagnation and blood stasis, according to a theory of traditional Chinese medicine. and (CRSR) is a classical herb pair that activates blood circulation to dissipate blood stasis. The purpose of this study was to explore the prevention and treatment effects of CRSR component compatibility on UL in rats. We randomly assigned adult female non-pregnant rats into three groups: a normal control (NC) group, a UL model group, and a CRSR treatment group. We administered to the UL and CRSR groups oral gavage diethylstilbestrol and injected them with progesterone (P) to establish UL for 5 weeks. The CRSR group received a CRSR medicinal solution after daily modeling. The uterus morphology of the UL group showed significantly more swelling than did that of the NC group, and we found no significant abnormalities in the morphology of the CRSR group. The pathological changes associated with UL were relieved in the CRSR group. CRSR improved the related parameters of the uterus and ovarian coefficients, significantly reducing the concentrations of P in the serum and the concentrations of estradiol, P, estrogen receptor, and P receptor in the uterus and ovary. In addition, CRSR significantly improved the abnormal blood conditions of UL, shown by decreases in plasma viscosity, the erythrocyte sedimentation rate equation value, and erythrocyte aggregation index. Therefore, CRSR component compatibility may prevent and cure UL through the above ways.
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http://dx.doi.org/10.3389/fpubh.2021.674357DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8195597PMC
June 2021

Deficiency of PPARγ in Bone Marrow Stromal Cells Does not Prevent High-Fat Diet-Induced Bone Deterioration in Mice.

J Nutr 2021 Sep;151(9):2697-2704

Department of Neuroscience and Regenerative Medicine, Augusta University, Augusta, GA, USA.

Background: Bone marrow osteoblasts and adipocytes are derived from a common mesenchymal stem cell and have a reciprocal relationship. Peroxisome proliferator-activated receptor gamma (PPARγ), a regulator for adipocyte differentiation, may be a potential target for reducing obesity and increasing bone mass.

Objectives: This study tested the hypothesis that bone-specific Pparg conditional knockout (cKO), via deletion of Pparg from bone marrow stromal cells (BMSC) using Osterix 1 (Osx1)-Cre, would prevent high-fat (HF) diet-induced bone deterioration in mice.

Methods: PPARγ cKO (PPARγfl/fl: Osx1-Cre) and floxed littermate control (PPARγfl/fl Osx1-Cre- ) mice that were 6 weeks old were randomly assigned to 4 groups (n = 12/group, 6 male and 6 female) and fed ad libitum with either a normal-fat (NF) purified diet (3.85 kcal/g; 10% energy as fat) or an HF diet (4.73 kcal/g; 45% energy as fat) for 6 mo. Bone structure, body composition, and serum bone-related cytokines were measured. Data were analyzed by 2-way ANOVA with Tukey post hoc comparison.

Results: The HF diet decreased the tibial and lumbar vertebrae trabecular bone volume/total volume (BV/TV) by 28% and 18%, respectively, compared to the NF diet (P < 0.01). PPARγ cKO mice had 23% lower body fat mass and 9% lower lean mass than control mice. PPARγ cKO mice had 41% greater tibial trabecular BV/TV compared to control mice. None of trabecular bone parameters at the second lumbar vertebra were affected by genotype. PPARγ cKO mice had decreased cortical thickness compared to control mice. PPARγ cKO mice had a 14% lower (P < 0.01) serum concentration of leptin and a 35% higher (P < 0.05) concentration of osteocalcin compared with control mice.

Conclusions: These data indicate that PPARγ has site-specific impacts on bone structures in mice and that knockout PPARγ in BMSC increased bone mass (BV/TV) in the tibia but not the lumbar vertebrae. PPARγ disruption in BMSC did not prevent HF diet-induced bone deterioration in mice.
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http://dx.doi.org/10.1093/jn/nxab173DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8417918PMC
September 2021

Inhibition of Fibroblast Activation in Uterine Leiomyoma by Components of and .

Front Public Health 2021 1;9:650022. Epub 2021 Mar 1.

Basic Medicine College, Chengdu University of Traditional Chinese Medicine, Chengdu, China.

The herbs and (RCRS) are often used in traditional Chinese medicine for the treatment of uterine leiomyoma (UL). The effectiveness of RCRS for the treatment of UL has been confirmed in our previous studies. This study aimed to investigate the molecular mechanism by which RCRS inhibits the activation of fibroblast activation protein (FAP) and prevents UL in rats. A Sprague Dawley (SD) rat model of UL was established via estrogen and progesterone load combined with external stimulation. Histological analyses, enzyme-linked immunosorbent assays, and western blotting were performed to evaluate the effect of RCRS on UL and elucidate its mechanism of action. Our data showed that the treatment of SD rats with RCRS significantly reduced the expression of extracellular matrix component collagen, FAP, and transforming growth factor beta (a FAP-activating factor) and the phosphorylation of the cell proliferation pathway-related signaling factors AKT/MEK/ERK. Our results suggest that RCRS is effective in the prevention and treatment of UL in rats, and RCRS may exert its functions by inhibiting the activation of tumor-associated fibroblasts and cell proliferation and by improving the tumor extracellular matrix.
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http://dx.doi.org/10.3389/fpubh.2021.650022DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7957009PMC
May 2021

Deletion of PPARγ in Mesenchymal Lineage Cells Protects Against Aging-Induced Cortical Bone Loss in Mice.

J Gerontol A Biol Sci Med Sci 2020 04;75(5):826-834

Center for Healthy Aging, Augusta University, Georgia.

Bone loss in aging is linked with chronic low-grade inflammation and the accumulation of marrowfat in animals and humans. Peroxisome proliferator-activated receptor gamma (PPARγ), an adipogenic regulator, plays key roles in these biological processes. However, studies of the roles of PPARγ in age-related bone loss and inflammation are lacking. We hypothesized that deletion of PPARγ in bone marrow mesenchymal lineage cells would reduce bone loss with aging, potentially through a reduction in fat-generated inflammatory responses and an increase in osteoblastic activity. In the present study, we show that mice deficient of PPARγ in Dermo1-expressing mesenchymal lineage cells (Dermo1-Cre:PPARγ fl/fl) have reduced fat mass and increased cortical bone thickness but that deficiency of PPARγ had limited effect on protection of trabecular bone with aging as demonstrated by dual-energy X-ray absorptiometry, µCT, and histomorphometric analyses. Conditional knockout of PPARγ reduced serum concentrations of adipokines, including adiponectin, resistin, and leptin, and reduced marrow stromal cell expression levels of inflammation-related genes. Inflammation genes involved in the interferon signaling pathway were reduced the most. These results demonstrate that disruption of the master adipogenic regulator, PPARγ, has a certain protective effect on aging-induced bone loss, suggesting that regulation of adipose function and modulation of interferon signaling are among the key mechanisms by which PPARγ regulates bone homeostasis during aging process.
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http://dx.doi.org/10.1093/gerona/glaa049DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7164529PMC
April 2020

Endogenous Glucocorticoid Signaling in the Regulation of Bone and Marrow Adiposity: Lessons from Metabolism and Cross Talk in Other Tissues.

Curr Osteoporos Rep 2019 12;17(6):438-445

Department of Cellular Biology and Anatomy, Medical College of Georgia, Augusta University, 1460 Laney Walker Blvd., CB1101, Augusta, GA, USA.

Purpose Of Review: The development of adiposity in the bone marrow, known as marrow adipose tissue (MAT), is often associated with musculoskeletal frailty. Glucocorticoids, which are a key component of the biological response to stress, affect both bone and MAT. These molecules signal through receptors such as the glucocorticoid receptor (GR), but the role of the GR in regulation of MAT is not yet clear from previous studies. The purpose of this review is to establish and determine the role of GR-mediated signaling in marrow adiposity by comparing and contrasting what is known against other energy-storing tissues like adipose tissue, liver, and muscle, to provide better insight into the regulation of MAT during times of metabolic stress (e.g., dietary challenges, aging).

Recent Findings: GR-mediated glucocorticoid signaling is critical for proper storage and utilization of lipids in cells such as adipocytes and hepatocytes and proteolysis in muscle, impacting whole-body composition, energy utilization, and homeostasis through a complex network of tissue cross talk between these systems. Loss of GR signaling in bone promotes increased MAT and decreased bone mass. GR-mediated signaling in the liver, adipose tissue, and muscle is critical for whole-body energy and metabolic homeostasis, and both similarities and differences in GR-mediated GC signaling in MAT as compared with these tissues are readily apparent. It is clear that GC-induced pathways work together through these tissues to affect systemic biology, and understanding the role of bone in these patterns of tissue cross talk may lead to a better understanding of MAT-bone biology that improves treatment strategies for frailty-associated diseases.
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http://dx.doi.org/10.1007/s11914-019-00554-6DOI Listing
December 2019

Amino acids as signaling molecules modulating bone turnover.

Bone 2018 10 27;115:15-24. Epub 2018 Feb 27.

Institute for Regenerative and Reparative Medicine, Medical College of Georgia, Augusta University, USA; Department of Neuroscience and Regenerative Medicine, Medical College of Georgia, Augusta University, USA; Department of Orthopaedic Surgery, Medical College of Georgia, Augusta University, USA; Department of Medicine, Medical College of Georgia, Augusta University, USA; Department of Cellular Biology and Anatomy, Medical College of Georgia, Augusta University, USA. Electronic address:

Except for the essential amino acids (AAs), much of the focus on adequate dietary protein intake has been on total nitrogen and caloric intake rather than AA composition. Recent data, however, demonstrate that "amino-acid sensing" can occur through either intracellular or extracellular nutrient-sensing mechanisms. In particular, members of the class 3 G-protein coupled receptor family, like the calcium-sensing receptor are known to preferentially bind specific AAs, which then modulate receptor activation by calcium ions and thus potentially impact bone turnover. In pursuing the possibility of direct nutrient effects on bone cells, we examined individual AA effects on osteoprogenitor/bone marrow stromal cells (BMSCs), a key target for bone anabolism. We demonstrate that BMSCs express both intracellular and extracellular nutrient sensing pathways and that AAs are required for BMSC survival. In addition, certain AA types, like members of the aromatic AAs, can potently stimulate increases in intracellular calcium and ERK phosphorylation/activation. Further, based on the in vitro data, we examined the effect of specific AAs on bone mass. To better evaluate the impact of specific AAs, we added these to a low-protein diet. Our data demonstrate that a low-protein diet itself is associated with a significant drop in bone mineral density (BMD) in the older mice, related, at least in part, to an increase in osteoclastic activity. This drop in BMD in mice on the low-protein diet is prevented by addition of AAs from the aromatic group. Taken together our data show that AAs function as specific and selective signaling molecules in bone cells.
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http://dx.doi.org/10.1016/j.bone.2018.02.028DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6110952PMC
October 2018

The role of GILZ in modulation of adaptive immunity in a murine model of myocardial infarction.

Exp Mol Pathol 2017 06 10;102(3):408-414. Epub 2017 May 10.

Department of Oral Biology, Augusta University, Augusta, GA 30912-1128, United States.

Myocardial infarction (MI) is associated with intense immune and inflammatory responses which contribute to tissue injury. Increasing evidence indicates that the glucocorticoid-induced leucine zipper (GILZ) protein suppresses immune and inflammatory responses. However, the status of and the role of GILZ in MI are not known. We tested the hypotheses that a) MI reduces cardiac GILZ associated with intense inflammation and cell death and b) intramyocardial GILZ delivery confers cardioprotection in association with increased Tregs and suppression of inflammation. Male Balb/C mice were subjected to MI or sham operation; the infarcted animals were subdivided to receive intramyocardial injections of PBS, GILZ overexpressing cells (GILZ) or their controls expressing the green fluorescent protein (GFP). Three hours after the procedures, hearts were procured for subsequent analyses. MI markedly reduced cardiac GILZ expression accompanied with a) increase in Th-17 cells (i.e., CD3CD4IL-17 BNP) but decrease in Tregs (i.e., CD3CD4FoxP3BNP), and b) disruption of mitochondrial membrane potential (ψ) associated with significant increases in apoptotic and necrotic cell death. While both GILZ and GFP returned the aforementioned parameters towards those of sham controls, these effects were most marked for mice receiving GILZ. Thus, GILZ markedly reduced Th-17 cells but increased Tregs and the anti-inflammatory cytokine, IL-10 positive cells accompanied with preservation of ψ and prevention of cell death. To our knowledge, this is the first report indicating an important role for GILZ in MI, in part via modulation of adaptive immune response, which raises the prospect of exogenous GILZ delivery as a novel cardioprotective modality.
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http://dx.doi.org/10.1016/j.yexmp.2017.05.002DOI Listing
June 2017

MicroRNA-183-5p Increases with Age in Bone-Derived Extracellular Vesicles, Suppresses Bone Marrow Stromal (Stem) Cell Proliferation, and Induces Stem Cell Senescence.

Tissue Eng Part A 2017 11 28;23(21-22):1231-1240. Epub 2017 Apr 28.

Department of Cellular Biology & Anatomy, Medical College of Georgia, Augusta University , Augusta, Georgia .

Microvesicle- and exosome-mediated transport of microRNAs (miRNAs) represents a novel cellular and molecular pathway for cell-cell communication. In this study, we tested the hypothesis that these extracellular vesicles (EVs) and their miRNAs might change with age, contributing to age-related stem cell dysfunction. EVs were isolated from the bone marrow interstitial fluid (supernatant) of young (3-4 months) and aged (24-28 months) mice to determine whether the size, concentration, and miRNA profile of EVs were altered with age in vivo. Results show that EVs isolated from bone marrow are CD63 and CD9 positive, and the concentration and size distribution of bone marrow EVs are similar between the young and aged mice. Bioanalyzer data indicate that EVs from both young and aged mice are highly enriched in miRNAs, and the miRNA profile of bone marrow EVs differs significantly between the young and aged mice. Specifically, the miR-183 cluster (miR-96/-182/-183) is highly expressed in aged EVs. In vitro assays demonstrate that aged EVs are endocytosed by primary bone marrow stromal cells (BMSCs), and these aged EVs inhibit the osteogenic differentiation of young BMSCs. Transfection of BMSCs with miR-183-5p mimic reduces cell proliferation and osteogenic differentiation, increases senescence, and decreases protein levels of the miR-183-5p target heme oxygenase-1 (Hmox1). In vitro assays utilizing HO-induced oxidative stress show that HO treatment of BMSCs increases the abundance of miR-183-5p in BMSC-derived EVs, and Amplex Red assays demonstrate that HO is elevated in the bone marrow microenvironment with age. Together, these data indicate that aging and oxidative stress can significantly alter the miRNA cargo of EVs in the bone marrow microenvironment, which may in turn play a role in stem cell senescence and osteogenic differentiation by reducing Hmox1 activity.
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http://dx.doi.org/10.1089/ten.TEA.2016.0525DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5689127PMC
November 2017

Analysis of the spleen proteome of chickens infected with reticuloendotheliosis virus.

Arch Virol 2017 May 17;162(5):1187-1199. Epub 2017 Jan 17.

State Key Laboratory of Veterinary Biotechnology, Harbin Veterinary Research Institute, The Chinese Academy of Agricultural Sciences, Harbin, 150001, People's Republic of China.

Infection with reticuloendotheliosis virus (REV), a gammaretrovirus in the family Retroviridae, can result in immunosuppression and subsequent increased susceptibility to secondary infections. In the present study, we identified differentially expressed proteins in the spleens of chickens infected with the REV-A HLJ07I strain, using two-dimensional gel electrophoresis on samples from time points coinciding with different phases of the REV life cycle. Differentially expressed proteins were identified using one-dimensional liquid chromatography electrospray ionization tandem mass spectrometry (1D LC ESI MS/MS). Comparative analysis of multiple gels revealed that the majority of changes occurred at early stages of infection. In total, 60 protein spots representing 28 host proteins were detected as either quantitatively (false discovery rate [FDR] ≤0.05 and fold change ≥2) or qualitatively differentially expressed at least once during different sampling points. The differentially expressed proteins identified in this study included antioxidants, molecular chaperones, cellular metabolism, formation of the cytoskeleton, signal transduction, cell proliferation and cellar aging. The present findings provide a basis for further studies to elucidate the role of these proteins in REV-host interactions. This could lead to a better understanding of REV infection mechanisms that cause immune suppression.
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http://dx.doi.org/10.1007/s00705-016-3180-5DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5387025PMC
May 2017

The status of glucocorticoid-induced leucine zipper protein in the salivary glands in Sjögren's syndrome: predictive and prognostic potentials.

EPMA J 2015 5;7. Epub 2016 Feb 5.

Department of Oral Biology, Dental College of Georgia, Augusta University, Augusta, GA 30912 USA ; Section of Plastic Surgery, Department of Surgery, Medical College of Georgia, Augusta University, Augusta, GA 30912 USA.

Background: We recently showed that an imbalance between the pro-inflammatory cytokine, interleukin (IL)-17, and the developmental endothelial locus-1 (Del-1) likely contributes to inflammation and salivary gland abnormalities in Sjögren's syndrome (SS). The glucocorticoid-induced leucine zipper (GILZ) protein is a pivotal player in mediating the anti-inflammatory effects of glucocorticoids. However, its status and role in salivary gland inflammation and dysfunction in SS are not established. Thus, we tested the hypothesis that SS is associated with reduced GILZ expression, thereby contributing to Del-1/Il-17 imbalance and inflammation in salivary glands.

Methods: We utilized the nonobese diabetic (NOD) mice, a model of SS-like disease as well as lower-lip biopsy samples of subjects without or with a diagnosis of SS in association with immunostaining studies. These studies were complemented with in vitro and flow-cytometry studies whereby interleukin (IL)-23-treated salivary gland cells were co-cultured with GILZ-expressing cells or control cells; IL-23 is known to increase generation of IL-17.

Results: Salivary glands of NOD mice displayed marked leukocyte infiltration and reduced GILZ expression in association with increased IL-17 but decreased Del-1 expression. A similar pattern was observed for lower-lip biopsy samples of SS than non-SS subjects. Further, IL-23-treated salivary gland cells displayed marked increase in IL-17 but reduced Del-1 positive cells which were reversed with co-culturing with GILZ-expressing cells but not control cells. Collectively, the results are suggestive of dysregulation of GILZ playing a role in inflammation and associated Del-1/Il-17 imbalance in SS.

Conclusions: Complementing our demonstration of Del-1/IL-17 imbalance in salivary glands in SS, the present study has established the relevance and significance of GILZ as a novel predictive and prognostic molecular fingerprint for SS. Thus, assessment of minor salivary gland GILZ expression, in conjunction with Del-1/IL-17 imbalance, could potentially offer a more sensitive approach to help with diagnosis of SS, at early stage of the disease, than that based on leukocyte infiltration. Future studies should establish whether treatment with GILZ ameliorates signs and symptoms of salivary malfunction of SS for which effective treatment options remain elusive.
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http://dx.doi.org/10.1186/s13167-016-0052-8DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4743166PMC
February 2016

Mesenchymal stem cell expression of SDF-1β synergizes with BMP-2 to augment cell-mediated healing of critical-sized mouse calvarial defects.

J Tissue Eng Regen Med 2017 06 31;11(6):1806-1819. Epub 2015 Jul 31.

Department of Oral Health Sciences, Medical University of South Carolina, Charleston, SC, USA.

Bone has the potential for spontaneous healing. This process, however, often fails in patients with comorbidities. Tissue engineering combining functional cells, biomaterials and osteoinductive cues may provide alternative treatment strategies. We have recently demonstrated that stromal cell-derived factor-1β (SDF-1β) works in concert with bone morphogenetic protein-2 (BMP-2) to potentiate osteogenic differentiation of bone marrow-derived mesenchymal stem/stromal cells (BMSCs). Here, we test the hypothesis that SDF-1β overexpressed in Tet-Off-SDF-1β BMSCs, delivered on acellular dermal matrix (ADM), synergistically augments BMP-2-induced healing of critical-sized mouse calvarial defects. BMSC therapies alone showed limited bone healing, which was increased with co-delivery of BMP-2. This was further enhanced in Tet-Off-SDF-1β BMSCs + BMP-2. Only limited BMSC retention on ADM constructs was observed after 4 weeks in vivo, which was increased with BMP-2 co-delivery. In vitro cell proliferation studies showed that supplementing BMP-2 to Tet-Off BMSCs significantly increased the cell number during the first 24 h. Consequently, the increased cell numbers decreased the detectable BMP-2 levels in the medium, but increased cell-associated BMP-2. The data suggest that SDF-1β provides synergistic effects supporting BMP-2-induced, BMSC-mediated bone formation and appears suitable for optimization of bone augmentation in combination therapy protocols. Copyright © 2015 John Wiley & Sons, Ltd.
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http://dx.doi.org/10.1002/term.2078DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4733586PMC
June 2017

Mesenchymal stem cell expression of stromal cell-derived factor-1β augments bone formation in a model of local regenerative therapy.

J Orthop Res 2015 Feb 28;33(2):174-84. Epub 2014 Oct 28.

Department of Biomedical Engineering, Case Western Reserve University, Cleveland, Ohio.

Bone has the potential for spontaneous healing. However, this process often fails in patients with co-morbidities requiring clinical intervention. Numerous studies have revealed that bone marrow-derived mesenchymal stem/stromal cells (BMSCs) hold great potential for regenerative therapies. Common problems include poor cell engraftment, which can be addressed by irradiation prior to transplantation. Increasing evidence suggests that stromal cell-derived factor-1 (SDF-1) is involved in bone formation. However, osteogenic contributions of the beta splice variant of SDF-1 (SDF-1β), which is highly expressed in bone, remain unclear. Using the tetracycline (Tet)-regulatory system we have shown that SDF-1β enhances BMSC osteogenic differentiation in vitro. Here we test the hypothesis that SDF-1β augments bone formation in vivo in a model of local BMSC transplantation following irradiation. We found that SDF-1β, expressed at high levels in Tet-Off-SDF-1β BMSCs, augments the cell-mediated therapeutic effects resulting in enhanced bone formation, as evidenced by ex vivo μCT and bone histomorphometry. The data demonstrate the specific contribution of SDF-1β to BMSC-mediated bone formation, and validate the feasibility of the Tet-Off technology to regulate SDF-1β expression in vivo. In conclusion, SDF-1β provides potent synergistic effects supporting BMSC-mediated bone formation and appears a suitable candidate for optimization of bone augmentation in translational protocols.
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http://dx.doi.org/10.1002/jor.22749DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4706461PMC
February 2015

Total body irradiation is permissive for mesenchymal stem cell-mediated new bone formation following local transplantation.

Tissue Eng Part A 2014 Dec;20(23-24):3212-27

1 Charlie Norwood VA Medical Center, Georgia Regents University , Augusta, Georgia .

Skeletal injury is a major clinical challenge accentuated by the decrease of bone marrow-derived mesenchymal stem/stromal cells (BMSCs) with age or disease. Numerous experimental and clinical studies have revealed that BMSCs hold great promise for regenerative therapies due to their direct osteogenic potential and indirect trophic/paracrine actions. Increasing evidence suggests that stromal cell-derived factor-1 (SDF-1) is involved in modulating the host response to the injury. Common problems with BMSC therapy include poor cell engraftment, which can be addressed by total body irradiation (TBI) prior to transplantation. In this study, we tested the hypothesis that direct tibial transplantation of BMSCs drives endogenous bone formation in a dose-dependent manner, which is enhanced by TBI, and investigated the potential role of SDF-1 in facilitating these events. We found that TBI is permissive for transplanted BMSCs to engraft and contribute to new bone formation. Bone marrow (BM) interstitial fluid analysis revealed no differences of SDF-1 splice variants in irradiated animals compared to controls, despite the increased mRNA and protein levels expressed in whole BM cells. This correlated with increased dipeptidyl peptidase IV activity and the failure to induce chemotaxis of BMSCs in vitro. We found increased mRNA expression levels of the major SDF-1-cleaving proteases in whole BM cells from irradiated animals suggesting distinct spatial differences within the BM in which SDF-1 may play different autocrine and paracrine signaling roles beyond the immediate cell surface microenvironment.
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http://dx.doi.org/10.1089/ten.TEA.2013.0663DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4259199PMC
December 2014

Effects of reticuloendotheliosis virus infection on cytokine production in SPF chickens.

PLoS One 2013 16;8(12):e83918. Epub 2013 Dec 16.

Division of Avian Infectious Diseases, State Key Laboratory of Veterinary Biotechnology, Harbin Veterinary Research Institute, The Chinese Academy of Agricultural Sciences, Harbin, China ; National Engineering Research Center of Veterinary Biologics, Harbin, China.

Infection with reticuloendotheliosis virus (REV), a gammaretrovirus in the Retroviridae family, can result in immunosuppression and subsequent increased susceptibility to secondary infections. The effects of REV infection on expression of mRNA for cytokine genes in chickens have not been completely elucidated. In this study, using multiplex branched DNA (bDNA) technology, we identified molecular mediators that participated in the regulation of the immune response during REV infection in chickens. Cytokine and chemokine mRNA expression levels were evaluated in the peripheral blood mononuclear cells (PBMCs). Expression levels of interleukin (IL)-4, IL-10, IL-13 and tumor necrosis factor (TNF)-α were significantly up-regulated while interferon (IFN)-α, IFN-β, IFN-γ, IL-1β, IL-2, IL-3, IL-15, IL-17F, IL-18 and colony-stimulating factor (CSF)-1 were markedly decreased in PBMCs at all stages of infection. Compared with controls, REV infected chickens showed greater expression levels of IL-8 in PBMCs 21 and 28 days post infection. In addition, REV regulates host immunity as a suppressor of T cell proliferative responses. The results in this study will help us to understand the host immune response to virus pathogens.
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http://journals.plos.org/plosone/article?id=10.1371/journal.pone.0083918PLOS
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3865284PMC
September 2014

Detection of infectious laryngotracheitis virus by real-time PCR in naturally and experimentally infected chickens.

PLoS One 2013 28;8(6):e67598. Epub 2013 Jun 28.

Division of Avian Infectious Diseases, State Key Laboratory of Veterinary Biotechnology, Harbin Veterinary Research Institute, The Chinese Academy of Agricultural Sciences, Harbin, China.

Infectious laryngotracheitis (ILT) is an acute, highly contagious upper-respiratory infectious disease of chickens. In this study, a real-time PCR method was developed for fast and accurate detection and quantitation of ILTV DNA of chickens experimentally infected with ILTV strain LJS09 and naturally infected chickens. The detection lower limit of the assay was 10 copies of DNA. There were no cross reactions with the DNA and RNA of infectious bursal disease virus, chicken anemia virus, reticuloendotheliosis virus, avian reovirus, Newcastle disease virus, and Marek's disease virus. The real-time PCR was reproducible as the coefficients of variation of reproducibility of the intra-assay and the inter-assay were less than 2%. The real-time PCR was used to detect the levels of the ILTV DNA in the tissues of specific pathogen free (SPF) chickens infected with ILTV at different times post infection. ILTV DNA was detected by real-time PCR in the heart, liver, spleen, lung, kidney, larynx, tongue, thymus, glandular stomach, duodenum, pancreatic gland, small intestine, large intestine, cecum, cecal tonsil, bursa of Fabricius, and brain of chickens in the infection group and the contact-exposure group. The sensitivity, specificity, and reproducibility of the ILTV real-time PCR assay revealed its suitability for detection and quantitation of ILTV in the samples from clinically and experimentally ILTV infected chickens.
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http://journals.plos.org/plosone/article?id=10.1371/journal.pone.0067598PLOS
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3695875PMC
April 2014

Stromal cell-derived factor-1β mediates cell survival through enhancing autophagy in bone marrow-derived mesenchymal stem cells.

PLoS One 2013 5;8(3):e58207. Epub 2013 Mar 5.

Charlie Norwood VA Medical Center, Augusta, Georgia, USA.

Bone marrow-derived mesenchymal stem/stromal cells (BMSCs) hold great potential for cell-based therapy, yet the therapeutic efficacy remains uncertain. Transplanted BMSCs often fail to engraft within the bone marrow (BM), in part due to the poor survival of donor cells in response to inflammatory reactions, hypoxia, oxidative stress, or nutrient starvation. Two basic cell processes, apoptosis and autophagy, could potentially be responsible for the impaired survival of transplanted BMSCs. However, the functional relationship between apoptosis and autophagy in BMSC homeostasis is complex and not well understood. The stromal cell-derived factor-1 (SDF-1)/CXC chemokine receptor 4 (CXCR4) signaling axis appears to be critical in maintaining proliferation and survival of BM stem cell populations through improving cell proliferation and survival in response to stress; however, the exact mechanisms remain unclear. We recently described novel genetically engineered Tet-Off-SDF-1β BMSCs, which over-express SDF-1β under tight doxycycline-control, thus providing an ideal model system to investigate the isolated effects of SDF-1β. In this study we tested the hypothesis that SDF-1β can mediate cell survival of BMSCs in vitro through increasing autophagy. We found that SDF-1β had no effect on BMSC proliferation; however, SDF-1β significantly protected genetically engineered BMSCs from H2O2-induced cell death through increasing autophagy and decreasing caspase-3-dependent apoptosis. Taken together, we provide novel evidence that the SDF-1/CXCR4 axis, specifically activated by the SDF-1β isoform, plays a critical role in regulating BMSC survival under oxidative stress through increasing autophagy.
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http://journals.plos.org/plosone/article?id=10.1371/journal.pone.0058207PLOS
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3589360PMC
December 2013

Avirulent Marek's disease virus type 1 strain 814 vectored vaccine expressing avian influenza (AI) virus H5 haemagglutinin induced better protection than turkey herpesvirus vectored AI vaccine.

PLoS One 2013 3;8(1):e53340. Epub 2013 Jan 3.

Division of Avian Infectious Diseases, State Key Laboratory of Veterinary Biotechnology, Harbin Veterinary Research Institute, The Chinese Academy of Agricultural Sciences, Harbin, China.

Background: Herpesvirus of turkey (HVT) as a vector to express the haemagglutinin (HA) of avian influenza virus (AIV) H5 was developed and its protection against lethal Marek's disease virus (MDV) and highly pathogenic AIV (HPAIV) challenges was evaluated previously. It is well-known that avirulemt MDV type 1 vaccines are more effective than HVT in prevention of lethal MDV infection. To further increase protective efficacy against HPAIV and lethal MDV, a recombinant MDV type 1 strain 814 was developed to express HA gene of HPAIV H5N1.

Methodology/principal Findings: A recombinant MDV-1 strain 814 expressing HA gene of HPAIV H5N1 virus A/goose/Guangdong/3/96 at the US2 site (rMDV-HA) was developed under the control of a human CMV immediate-early promoter. The HA expression in the rMDV-HA was tested by immunofluorescence and Western blot analyses, and in vitro and in vivo growth properties of rMDV-HA were also analyzed. Furthermore, we evaluated and compared the protective immunity of rMDV-HA and previously constructed rHVT-HA against HPAIV and lethal MDV. Vaccination of chickens with rMDV-HA induced 80% protection against HPAIV, which was better than the protection rate by rHVT-HA (66.7%). In the animal study with MDV challenge, chickens immunized with rMDV-HA were completely protected against virulent MDV strain J-1 whereas rHVT-HA only induced 80% protection with the same challenge dose.

Conclusions/significance: The rMDV-HA vaccine was more effective than rHVT-HA vaccine for protection against lethal MDV and HPAIV challenges. Therefore, avirulent MDV type 1 vaccine is a better vector than HVT for development of a recombinant live virus vaccine against virulent MDV and HPAIV in poultry.
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http://journals.plos.org/plosone/article?id=10.1371/journal.pone.0053340PLOS
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3536743PMC
August 2013

Identification of a conserved B-cell epitope on reticuloendotheliosis virus envelope protein by screening a phage-displayed random peptide library.

PLoS One 2012 21;7(11):e49842. Epub 2012 Nov 21.

Division of Avian Infectious Diseases, State Key Laboratory of Veterinary Biotechnology, Harbin Veterinary Research Institute, The Chinese Academy of Agricultural Sciences, Harbin, P. R. China.

Background: The gp90 protein of avian reticuloendotheliosis-associated virus (REV-A) is an important envelope glycoprotein, which is responsible for inducing protective antibody immune responses in animals. B-cell epitopes on the gp90 protein of REV have not been well studied and reported.

Methods And Results: This study describes the identification of a linear B-cell epitope on the gp90 protein by screening a phage-displayed 12-mer random peptide library with the neutralizing monoclonal antibody (mAb) A9E8 directed against the gp90. The mAb A9E8 recognized phages displaying peptides with the consensus motif SVQYHPL. Amino acid sequence of the motif exactly matched (213)SVQYHPL(219) of the gp90. Further identification of the displayed B cell epitope was conducted using a set of truncated peptides expressed as GST fusion proteins and the Western blot results indicated that (213)SVQYHPL(219) was the minimal determinant of the linear B cell epitope recognized by the mAb A9E8. Moreover, an eight amino acid peptide SVQYHPLA was proven to be the minimal unit of the epitope with the maximal binding activity to mAb A9E8. The REV-A-positive chicken serum reacted with the minimal linear epitopes in Western blot, revealing the importance of the eight amino acids of the epitope in antibody-epitope binding activity. Furthermore, we found that the epitope is a common motif shared among REV-A and other members of REV group.

Conclusions And Significance: We identified (213)SVQYHPL(219) as a gp90-specific linear B-cell epitope recognized by the neutralizing mAb A9E8. The results in this study may have potential applications in development of diagnostic techniques and epitope-based marker vaccines against REV-A and other viruses of the REV group.
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http://journals.plos.org/plosone/article?id=10.1371/journal.pone.0049842PLOS
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3504085PMC
May 2013

Effects of the activin A-myostatin-follistatin system on aging bone and muscle progenitor cells.

Exp Gerontol 2013 Feb 21;48(2):290-7. Epub 2012 Nov 21.

Georgia Health Sciences University, Augusta, GA 30912, USA.

The activin A-myostatin-follistatin system is thought to play an important role in the regulation of muscle and bone mass throughout growth, development, and aging; however, the effects of these ligands on progenitor cell proliferation and differentiation in muscle and bone are not well understood. In addition, age-associated changes in the relative expression of these factors in musculoskeletal tissues have not been described. We therefore examined changes in protein levels of activin A, follistatin, and myostatin (GDF-8) in both muscle and bone with age in C57BL6 mice using ELISA. We then investigated the effects of activin A, myostatin and follistatin on the proliferation and differentiation of primary myoblasts and mouse bone marrow stromal cells (BMSCs) in vitro. Myostatin levels and the myostatin:follistatin ratio increased with age in the primarily slow-twitch mouse soleus muscle, whereas the pattern was reversed with age in the fast-twitch extensor digitorum longus muscle. Myostatin levels and the myostatin:follistatin ratio increased significantly (+75%) in mouse bone marrow with age, as did activin A levels (+17%). Follistatin increased the proliferation of primary myoblasts from both young and aged mice, whereas myostatin increased proliferation of younger myoblasts but decreased proliferation of older myoblasts. Myostatin reduced proliferation of both young and aged BMSCs in a dose-dependent fashion, and activin A increased mineralization in both young and aged BMSCs. Together these data suggest that aging in mice is accompanied by changes in the expression of activin A and myostatin, as well as changes in the response of bone and muscle progenitor cells to these factors. Myostatin appears to play a particularly important role in the impaired proliferative capacity of muscle and bone progenitor cells from aged mice.
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http://dx.doi.org/10.1016/j.exger.2012.11.004DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3678732PMC
February 2013

Stromal cell-derived factor-1β potentiates bone morphogenetic protein-2-stimulated osteoinduction of genetically engineered bone marrow-derived mesenchymal stem cells in vitro.

Tissue Eng Part A 2013 Jan 21;19(1-2):1-13. Epub 2012 Aug 21.

Charlie Norwood Veterans Affairs Medical Center, Augusta, Georgia, USA.

Skeletal injuries are among the most prevalent clinical problems and bone marrow-derived mesenchymal stem/stromal cells (BMSCs) have successfully been used for the treatment thereof. Stromal cell-derived factor-1 (SDF-1; CXCL12) is a member of the CXC chemokine family with multiple splice variants. The two most abundant variants, SDF-1α and SDF-1β, share identical amino acid sequences, except for four additional amino acids at the C-terminus of SDF-1β, which may mediate surface stabilization via glycosaminoglycans and protect SDF-1β from proteolytic cleavage, rendering it twice as potent as SDF-1α. Increasing evidence suggests that SDF-1 is involved in bone formation through regulation of recruitment, engraftment, proliferation, and differentiation of stem/progenitor cells. The underlying molecular mechanisms, however, have not yet been fully elucidated. In this study, we tested the hypothesis that SDF-1β can potentiate bone morphogenetic protein-2 (BMP-2)-stimulated osteogenic differentiation and chemotaxis of BMSCs in vitro. Utilizing retrovirus-mediated gene transfer to generate novel Tet-Off-SDF-1β BMSCs, we found that conditional SDF-1β expression is tightly regulated by doxycycline in a dose-dependent and temporal fashion, leading to significantly increased SDF-1β mRNA and protein levels. In addition, SDF-1β was found to enhance BMP-2-stimulated mineralization, mRNA and protein expression of key osteogenic markers, and regulate BMP-2 signal transduction via extracellular signal-regulated kinases 1/2 (Erk1/2) phosphorylation in genetically engineered BMSCs in vitro. We also showed that SDF-1β promotes the migratory response of CXC chemokine receptor 4 (CXCR4)-expressing BMSCs in vitro. Taken together, these data support that SDF-1β can play an important role in BMP-2-stimulated osteogenic differentiation of BMSCs and may exert its biological activity in both an autocrine and paracrine fashion.
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http://dx.doi.org/10.1089/ten.TEA.2012.0085DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3530941PMC
January 2013

A comparative study of bone marrow mesenchymal stem cell functionality in C57BL and mdx mice.

Neurosci Lett 2012 Aug 3;523(2):139-44. Epub 2012 Jul 3.

Department of Neurology, The First Affiliated Hospital, Sun Yat-sen University, No. 58 Zhongshan Road 2, Guangzhou 510080, China.

Patients with DMD have low bone mass and a high incidence of fractures, but the cellular and molecular mechanisms underlying this pathological condition are unknown. Because bone marrow mesenchymal stem cells (BMSCs) are the progenitors of bone-forming osteoblasts, we hypothesized that DMD leads to dysfunction in the differentiation of BMSCs. We isolated BMSCs from C57BL control and mdx mutant mice, a well-established mouse model of DMD, and compared their abilities of proliferation, differentiation, and the expression of lineage-specific genes. Results showed that the proliferation and osteogenic and myogenic differentiation of BMSCs from mdx mice were significantly lower than those from C57BL mice. Because mutations in dystrophin gene cause DMD, our results demonstrate that dystrophin deficiency leads to dysfunction in the differentiation and proliferation of BMSCs.
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http://dx.doi.org/10.1016/j.neulet.2012.06.061DOI Listing
August 2012

Development of a rapid and specific loop-mediated isothermal amplification detection method that targets Marek's disease virus meq gene.

J Virol Methods 2012 Aug 30;183(2):196-200. Epub 2012 Apr 30.

Division of Avian Infectious Diseases, State Key Laboratory of Veterinary Biotechnology, Harbin Veterinary Research Institute, The Chinese Academy of Agricultural Sciences, Harbin 150001, China.

A rapid, sensitive and specific loop-mediated isothermal amplification (LAMP) method was developed and evaluated for the detection of Marek's disease virus (MDV) by amplification of conserved MDV meq gene sequences. LAMP is an innovative technique that allows the rapid detection of targeted nucleic acid sequences under isothermal conditions without the need for complex instrumentation. In this study, meq gene sequences were amplified successfully from different MDV strains by LAMP within 60min and no cross-reactivity was observed in a panel of related viruses that were associated with diseases of chickens. The detection limit of LAMP was 3.2 copies/million cells compared with 320 copies/million cells required for conventional PCR. Positive detection rates were assessed using either LAMP or PCR by examination of feather follicles that were collected from chickens infected experimentally with either strain J-1 (n=20) or strain Md5 (n=17), In addition to these samples, three isolates that were suspected to have been infected in the clinic were also tested. Results showed that the positive detection rate for LAMP was 95% (38/40), compared with 87.5% (35/40) and 90% (38/40) for strains J-1 and Md5 by PCR, respectively. These results indicated that the LAMP assay was more sensitive, rapid and specific than conventional PCR for the detection of MDV. This easy-to-perform technique will be useful for the detection of MDV and will aid in the establishment of disease control protocols.
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http://dx.doi.org/10.1016/j.jviromet.2012.04.014DOI Listing
August 2012

Preparation and immunological effectiveness of a swine influenza DNA vaccine encapsulated in chitosan nanoparticles.

Vaccine 2011 Nov 21;29(47):8549-56. Epub 2011 Sep 21.

College of Life Science, Heilongjiang University, Harbin, China.

Preparation conditions of a DNA vaccine against swine influenza encapsulated in chitosan nanoparticles were determined. The nanoparticles were prepared according to a complex coacervation method using chitosan as a biodegradable matrix forming polymer. Under the preparation conditions, chitosan nanoparticles containing the DNA vaccine were produced with good morphology, high encapsulation rate and high stability. Transfection test indicated that the vaccine could be expressed as an antigen in cells, and maintained good bioactivity. In addition, better immune responses of mice immunized with the chitosan nanoparticles containing the DNA vaccine were induced and prolonged release of the plasmid DNA was achieved compared to the DNA vaccine alone. These results laid a foundation for further development of DNA vaccines in nanoparticles before ultimate industrial application.
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http://dx.doi.org/10.1016/j.vaccine.2011.09.029DOI Listing
November 2011

Expression of HA of HPAI H5N1 virus at US2 gene insertion site of turkey herpesvirus induced better protection than that at US10 gene insertion site.

PLoS One 2011 27;6(7):e22549. Epub 2011 Jul 27.

Division of Avian Infectious Diseases, State Key Laboratory of Veterinary Biotechnology, Harbin Veterinary Research Institute, The Chinese Academy of Agricultural Sciences, Harbin, China.

Herpesvirus of turkey (HVT) is being widely used as a vector for development of recombinant vaccines and US2 and US10 genes are often chosen as insertion sites for targeted gene expression. However, the different effects of the two genes for generation of recombinant HVT vaccines were unknown. In order to compare the effects of inserted genes in the two sites on the efficacy of the recombinant vaccines, host-protective haemagglutinin (HA) gene of the highly pathogenic avian influenza virus (HPAIV) H5N1 was inserted into either US2 or US10 gene locus of the HVT. The resulting US2 (rHVT-US2-HA) or US10 (rHVT-US10-HA) recombinant HVT viruses were used to infect chicken embryo fibroblasts. Plaques and the growth kinetics of rHVT-US2-HA-infected chicken embryo fibroblasts were similar to those of parental HVT whereas rHVT-US10-HA infected chicken embryo fibroblasts had different growth kinetics and plaque formation. The viremia levels in rHVT-US10-HA virus-infected chickens were significantly lower than those of rHVT-US2-HA group on 28 days post infection. The vaccine efficacy of the two recombinant viruses against H5N1 HPAIV and virulent Marek's disease virus was also evaluated in 1-day-old vaccinated chickens. rHVT-US2-HA-vaccinated chickens were better protected with reduced mortality than rHVT-US10-HA-vaccinated animals following HPAIV challenge. Furthermore, the overall hemaglutination inhibition antibody titers of rHVT-US2-HA-vaccinated chickens were higher than those of rHVT-US10-HA-vaccinated chickens. Protection levels against Marek's disease virus challenge following vaccination with either rHVT-US2-HA or rHVT-US10-HA, however, were similar to those of the parental HVT virus. These results, for the first time, indicate that US2 gene provides a favorable foreign gene insertion site for generation of recombinant HVT vaccines.
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http://journals.plos.org/plosone/article?id=10.1371/journal.pone.0022549PLOS
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3144902PMC
December 2011

Monitoring bone marrow-originated mesenchymal stem cell traffic to myocardial infarction sites using magnetic resonance imaging.

Magn Reson Med 2011 May 1;65(5):1430-6. Epub 2011 Feb 1.

Department of Radiology, Small Animal Imaging, Medical College of Georgia, Augusta, Georgia 30912, USA.

How stem cells promote myocardial repair in myocardial infarction (MI) is not well understood. The purpose of this study was to noninvasively monitor and quantify mesenchymal stem cells (MSC) from bone marrow to MI sites using magnetic resonance imaging (MRI). MSC were dual-labeled with an enhanced green fluorescent protein and micrometer-sized iron oxide particles prior to intra-bone marrow transplantation into the tibial medullary space of C57Bl/6 mice. Micrometer-sized iron oxide particles labeling caused signal attenuation in T(2)*-weighted MRI and thus allowed noninvasive cell tracking. Longitudinal MRI demonstrated MSC infiltration into MI sites over time. Fluorescence from both micrometer-sized iron oxide particles and enhanced green fluorescent protein in histology validated the presence of dual-labeled cells at MI sites. This study demonstrated that MSC traffic to MI sites can be noninvasively monitored in MRI by labeling cells with micrometer-sized iron oxide particles. The dual-labeled MSC at MI sites maintained their capability of proliferation and differentiation. The dual-labeling, intra-bone marrow transplantation, and MRI cell tracking provided a unique approach for investigating stem cells' roles in the post-MI healing process. This technique can potentially be applied to monitor possible effects on stem cell mobilization caused by given treatment strategies.
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http://dx.doi.org/10.1002/mrm.22735DOI Listing
May 2011

ACTH protects against glucocorticoid-induced osteonecrosis of bone.

Proc Natl Acad Sci U S A 2010 May 26;107(19):8782-7. Epub 2010 Apr 26.

The Mount Sinai Bone Program, Mount Sinai School of Medicine, New York, NY 10029, USA.

We report that adrenocorticotropic hormone (ACTH) protects against osteonecrosis of the femoral head induced by depot methylprednisolone acetate (depomedrol). This therapeutic response likely arises from enhanced osteoblastic support and the stimulation of VEGF by ACTH; the latter is largely responsible for maintaining the fine vascular network that surrounds highly remodeling bone. We suggest examining the efficacy of ACTH in preventing human osteonecrosis, a devastating complication of glucocorticoid therapy.
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http://dx.doi.org/10.1073/pnas.0912176107DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC2889316PMC
May 2010

[Culture optimization for protein expression in Escherichia coli].

Wei Sheng Wu Xue Bao 2009 Jan;49(1):128-34

Division of Avian Infectious Diseases, National Key Laboratory of Veterinary Biotechnology, Harbin Veterinary Research Institute, Chinese Academy of Agricultural Sciences, Harbin 150001, China.

Objective: We developed a new culture-complex auto-inducing media (CAI) for heterogenous protein expression in Escherichia coli.

Methods: To test expression efficiency in the CAI, we constructed seven different plasmids named p-1, p-2, p-3, p-4, p-5, p-6 and p-7. These plasmids were transformed into E. coli BL21, then expressed in both Luria-Bertani madia LB and CAI. To improve the expression level even more, we analyzed the composition of the CAI and optimized the culture.

Results: The expression levels of seven fusion proteins in CAI were four times higher than those in Luria-Bertani. Through a series of changes we formed a new optimized culture (CAI-4).

Conclusion: Comparing to the expression levels of these fusion proteins (P-1, P-2, P-3) in CAI, the expression levels of fusion proteins in CAI-4 increased 2-fold.
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January 2009
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