Publications by authors named "Manoj Bhaskaran"

23 Publications

  • Page 1 of 1

Scientific and Regulatory Policy Committee Best Practices: Documentation of Sexual Maturity by Microscopic Evaluation in Nonclinical Safety Studies.

Toxicol Pathol 2021 Mar 4:192623321990631. Epub 2021 Mar 4.

510456Idorsia Pharmaceuticals Limited, Allschwil, Switzerland.

The sexual maturity status of animals in nonclinical safety studies can have a significant impact on the microscopic assessment of the reproductive system, the interpretation of potential test article-related findings, and ultimately the assessment of potential risk to humans. However, the assessment and documentation of sexual maturity for animals in nonclinical safety studies is not conducted in a consistent manner across the pharmaceutical and chemical industries. The Scientific and Regulatory Policy Committee of the Society of Toxicologic Pathology convened an international working group of pathologists and nonclinical safety scientists with expertise in the reproductive system, pathology nomenclature, and Standard for Exchange of Nonclinical Data requirements. This article describes the best practices for documentation of the light microscopic assessment of sexual maturity in males and females for both rodent and nonrodent nonclinical safety studies. In addition, a review of the microscopic features of the immature, peripubertal, and mature male and female reproductive system and general considerations for study types and reporting are provided to aid the study pathologist tasked with documentation of sexual maturity.
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http://dx.doi.org/10.1177/0192623321990631DOI Listing
March 2021

The genome of the American groundhog, .

F1000Res 2020 16;9:1137. Epub 2020 Sep 16.

Department of Biomedical Engineering, Johns Hopkins University, Baltimore, MD, 21211, USA.

We sequenced the genome of the North American groundhog, , also known as the woodchuck. Our sequencing strategy included a combination of short, high-quality Illumina reads plus long reads generated by both Pacific Biosciences and Oxford Nanopore instruments. Assembly of the combined data produced a genome of 2.74 Gbp in total length, with an N50 contig size of 1,094,236 bp. To annotate the genome, we mapped the genes from another genome and from the closely related Alpine marmot, , onto our assembly, resulting in 20,559 annotated protein-coding genes and 28,135 transcripts. The genome assembly and annotation are available in GenBank under BioProject PRJNA587092.
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http://dx.doi.org/10.12688/f1000research.25970.1DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7682491PMC
September 2020

Reconstruction of Craniofacial Bone Defects with Autologous Human Bone Marrow Stem Cells and Autogenous Bone Grafts: A Case Report with Review of Literature.

J Pharm Bioallied Sci 2020 Aug 28;12(Suppl 1):S394-S398. Epub 2020 Aug 28.

Department of Oral and Maxillofacial Surgery, Sree Anjaneya Institute of Dental Sciences, Calicut, Kerala, India.

Reconstruction of craniofacial bony defects has always been a challenging task for the surgeons over the years. The science of reconstructing such defects is of at most importance to craniofacial and plastic surgeons due to its relevance in facial aesthetics function as well as prerequisite procedure for continuing other surgical procedures. The main goal of the reconstruction of the craniofacial defects is to reduce the morbidity by restoring the facial form and aesthetics, as well as a good function of the facial structures by achieving a reasonable occlusion and articulation. Although significant improvements have occurred during the last few decades, challenges still exist as to what type of reconstruction to be carried out with regard to techniques and the type and quality of materials of choice to be used. As decades progressed, the advancement in surgical techniques and the variety of reconstruction methods have definitely improved the quality of life. This article reviews the method of bony reconstruction of craniofacial defects using autologous human bone marrow stem cells and autologous bone grafts and its modification, which includes much recent tissue engineering techniques and regenerative medicine, thereby replacing older techniques by biological substitutes, which can restore improve and maintain orofacial function and aesthetics.
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http://dx.doi.org/10.4103/jpbs.JPBS_116_20DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7595491PMC
August 2020

Comparison of Efficacy of Lycopene and Lycopene-Hyaluronidase Combination in the Treatment of Oral Submucous Fibrosis.

J Pharm Bioallied Sci 2019 May;11(Suppl 2):S260-S264

Department of Oral and Maxillofacial Surgery, Sree Anjaneya Institute of Dental Sciences, Calicut, Kerala, India.

Background: Oral submucous fibrosis (OSMF) is a chronic progressively scarring disease of the oral cavity. Lycopene is a powerful antioxidant obtained from tomatoes and has the highest singlet oxygen quenching capacity and a high capacity of quenching other free radicals among dietary carotenoids. Hyaluronidase is a substance prepared from the testes and semen of mammals that modifies the permeability of connective tissue through the hydrolysis of hyaluronic acid.

Objective: To evaluate the efficacy of lycopene and lycopene-hyaluronidase combination, and to compare the efficacy of lycopene and lycopene-hyaluronidase combination in the treatment of OSMF.

Study Design: The study consisted of 45 patients with OSMF divided into three equal groups. Patients in Group A were given Lycored 16 mg daily in two equally divided doses for 3 months. Patients in Group B were given LycoRed along with hyaluronidase intralesional injection of 1500 IU twice weekly for 3 months. Patients in Group C were given placebo capsules. Patients were evaluated after 3 months. The following parameters were recorded: mouth opening, visual inspection, palpatory findings, and burning sensation.

Results: There was statistically significant change in mouth opening and burning sensation for lycopene and lycopene-hyaluronidase combination than in the placebo group in the treatment of OSMF, but the lycopene-hyaluronidase combination did not show any statistically significant change when compared with lycopene alone.

Conclusion: Lycopene appears to be a very promising antioxidant in the management of oral submucous fibrosis, both in clinical and symptomatic improvement.
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http://dx.doi.org/10.4103/JPBS.JPBS_6_19DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6555355PMC
May 2019

Immunotherapy in Oral Cancer.

J Pharm Bioallied Sci 2019 May;11(Suppl 2):S107-S111

Department of Oral and Maxillofacial Surgery, Sree Anjaneya Institute of Dental Sciences, Calicut, Kerala, India.

Immunotherapy is one of the newer entities which is promising, at least can be very much helpful as an adjuvant therapy. This newer modality of the treatment in the field of cancer treatment may be the fourth pillar supporting surgery, chemotherapy, and radiotherapy. Careful selection of patient is the key for success of immunotherapy, which is based on patient's immunological contexture. This review aimed to present the fundamental aspects of tumor immunity and immunotherapy, focused on oral squamous cell carcinoma.
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http://dx.doi.org/10.4103/JPBS.JPBS_31_19DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6555318PMC
May 2019

Sustained Klotho delivery reduces serum phosphate in a model of diabetic nephropathy.

J Appl Physiol (1985) 2019 04 3;126(4):854-862. Epub 2019 Jan 3.

Division of Molecular Genetics and Gene Therapy, Department of Medical and Molecular Genetics, Indiana University School of Medicine , Indianapolis, Indiana.

Diabetic nephropathy (DN) is a primary cause of end-stage renal disease and is becoming more prevalent because of the global rise in type 2 diabetes. A model of DN, the db/db uninephrectomized ( db/db-uni) mouse, is characterized by obesity, as well as compromised renal function. This model also manifests defects in mineral metabolism common in DN, including hyperphosphatemia, which leads to severe endocrine disease. The FGF23 coreceptor, α-Klotho, circulates as a soluble, cleaved form (cKL) and may directly influence phosphate handling. Our study sought to test the effects of cKL on mineral metabolism in db/db-uni mice. Mice were placed into either mild or moderate disease groups on the basis of the albumin-to-creatinine ratio (ACR). Body weights of db/db-uni mice were significantly greater across the study compared with lean controls regardless of disease severity. Adeno-associated cKL administration was associated with increased serum Klotho, intact, bioactive FGF23 (iFGF23), and COOH-terminal fragments of FGF23 ( P < 0.05). Blood urea nitrogen was improved after cKL administration, and cKL corrected hyperphosphatemia in the high- and low-ACR db/db-uni groups. Interestingly, 2 wk after cKL delivery, blood glucose levels were significantly reduced in db/db-uni mice with high ACR ( P < 0.05). Interestingly, several genes associated with stabilizing active iFGF23 were also increased in the osteoblastic UMR-106 cell line with cKL treatment. In summary, delivery of cKL to a model of DN normalized blood phosphate levels regardless of disease severity, supporting the concept that targeting cKL-affected pathways could provide future therapeutic avenues in DN. NEW & NOTEWORTHY In this work, systemic and continuous delivery of the "soluble" or "cleaved" form of the FGF23 coreceptor α-Klotho (cKL) via adeno-associated virus to a rodent model of diabetic nephropathy (DN), the db/db uninephrectomized mouse, normalized blood phosphate levels regardless of disease severity. This work supports the concept that targeting cKL-affected pathways could provide future therapeutic avenues for the severe mineral metabolism defects associated with DN.
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http://dx.doi.org/10.1152/japplphysiol.00838.2018DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6485689PMC
April 2019

Knowledge Analysis of Pit and Fissure Sealants among the Dental Students of South India.

J Int Soc Prev Community Dent 2018 Nov-Dec;8(6):508-512. Epub 2018 Nov 29.

Department of Oral and Maxillofacial Surgery, Sree Anjaneya Institute of Dental Science, Calicut, Kerala, India.

Aim: This study aimed to assess the level of knowledge and attitudes for pit and fissure sealants among undergraduate Indian dental students.

Materials And Methods: A modified questionnaire consisting of 24 items was distributed to 280 undergraduate dental students comprising males and females of different years at MNR Dental College, Sangareddy, India. Descriptive statistics and Chi-square/Fisher's exact tests were used for statistical analysis. The data were computationally tested using Statistical Package for the Social Sciences (SPSS) Version 20, IBM SPSS Statistics software for Windows, Armonk, NY, USA.

Results: With the response rate at 100%, most of the respondents, i.e., 70.4%, were females and the remaining 29.6% were male. Regarding the level of study, 20.8% were in 3 year, 43.8% in the 4 year, and 16.8% were in 5 year (internship). The respondents showed a reasonable level of knowledge about sealants, mostly being good with the theoretical concepts of the sealants. On the other hand, respondents showed insufficient knowledge about sealants in the clinical practice.

Conclusion: Although a high proportion of undergraduate dental students showed adequate knowledge about dental sealants, there is a lag in putting that knowledge into work during the clinical practice. These findings suggest an urgent need of dental schools to include and/or update their curriculum regarding fissure sealants to reflect modern dental education that concentrates on evidence-based practice in pediatric dentistry and improve the dental health among the future generations by reducing the incidence of caries.
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http://dx.doi.org/10.4103/jispcd.JISPCD_238_18DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6280565PMC
November 2018

Pancreatic Effects of a Bruton's Tyrosine Kinase Small-molecule Inhibitor in Rats Are Strain-dependent.

Toxicol Pathol 2018 06 26;46(4):460-472. Epub 2018 Apr 26.

1 Eli Lilly and Company, Indianapolis, Indiana, USA.

Inhibitors of Bruton's tyrosine kinase (BTK) are under development as potential therapies for various autoimmune diseases. In repeat-dose toxicity studies, small-molecule BTK inhibitors (BTKi) have been reported to cause a constellation of histologic effects at the pancreatic endocrine-exocrine interface in male rats; however, similar findings were not reported in other species. Since the BTKi-induced pancreatic effect is morphologically similar to well-documented spontaneous changes (predominantly characterized by insular/peri-insular hemorrhage, pigment deposition, chronic inflammation, and fibrosis) that are known to vary by rat strain, we investigated potential strain-dependent differences in the pancreatic effects of a small-molecule BTKi, LY3337641. Following 13 weeks of LY3337641 treatment, Crl:CD(SD) rats were most sensitive, Crl:WI(Han) rats were of intermediate sensitivity, and Hsd:SD rats were least sensitive. These strain differences appear to be related to differences in rate of weight gain across strains and sexes; however, a definitive mechanism was not determined. This study demonstrated that BTKi-induced pancreatic effects were highly dependent on rat strain and correlated with differences in the incidence and severity of the spontaneous background change. When considered with the lack of pancreas effects in nonrat species, these changes in rats are unlikely predictive of similar changes in humans administered a BTK inhibitor.
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http://dx.doi.org/10.1177/0192623318770163DOI Listing
June 2018

Chronic Hyperphosphatemia and Vascular Calcification Are Reduced by Stable Delivery of Soluble Klotho.

J Am Soc Nephrol 2017 Apr 11;28(4):1162-1174. Epub 2016 Nov 11.

Department of Medical and Molecular Genetics, Division of Molecular Genetics and Gene Therapy, Indiana University School of Medicine, Indianapolis, Indiana;

Klotho (KL) regulates mineral metabolism, and diseases associated with KL deficiency are characterized by hyperphosphatemia and vascular calcification (VC). KL is expressed as a membrane-bound protein (mKL) and recognized as the coreceptor for fibroblast growth factor-23 (FGF23) and a circulating soluble form (cKL) created by endoproteolytic cleavage of mKL. The functions of cKL with regard to phosphate metabolism are unclear. We tested the ability of cKL to regulate pathways and phenotypes associated with hyperphosphatemia in a mouse model of CKD-mineral bone disorder and α-null mice. Stable delivery of adeno-associated virus (AAV) expressing cKL to diabetic endothelial nitric oxide synthase-deficient mice or α-null mice reduced serum phosphate levels. Acute injection of recombinant cKL downregulated the renal sodium-phosphate cotransporter Npt2a in α-null mice supporting direct actions of cKL in the absence of mKL. α-null mice with sustained AAV-cKL expression had a 74%-78% reduction in aorta mineral content and a 72%-77% reduction in mineral volume compared with control-treated counterparts (<0.01). Treatment of UMR-106 osteoblastic cells with cKL + FGF23 increased the phosphorylation of extracellular signal-regulated kinase 1/2 and induced Fgf23 expression. CRISPR/Cas9-mediated deletion of fibroblast growth factor receptor 1 (FGFR1) or pretreatment with inhibitors of mitogen-activated kinase kinase 1 or FGFR ablated these responses. In summary, sustained cKL treatment reduced hyperphosphatemia in a mouse model of CKD-mineral bone disorder, and it reduced hyperphosphatemia and prevented VC in mice without endogenous KL. Furthermore, cKL stimulated Fgf23 in an FGFR1-dependent manner in bone cells. Collectively, these findings indicate that cKL has mKL-independent activity and suggest the potential for enhancing cKL activity in diseases of hyperphosphatemia with associated VC.
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http://dx.doi.org/10.1681/ASN.2015111266DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5373441PMC
April 2017

Clinical and Pathological Findings Associated with Aerosol Exposure of Macaques to Ricin Toxin.

Toxins (Basel) 2015 Jun 9;7(6):2121-33. Epub 2015 Jun 9.

Divisions of Pathology, Veterinary Medicine, and Microbiology, Tulane National Primate Research Center, 18703 Three Rivers Road, Covington, LA 70433, USA.

Ricin is a potential bioweapon that could be used against civilian and military personnel. Aerosol exposure is the most likely route of contact to ricin toxin that will result in the most severe toxicity. Early recognition of ricin exposure is essential if specific antidotes are to be applied. Initial diagnosis will most likely be syndromic, i.e., fitting clinical and laboratory signs into a pattern which then will guide the choice of more specific diagnostic assays and therapeutic interventions. We have studied the pathology of ricin toxin in rhesus macaques exposed to lethal and sublethal ricin aerosols. Animals exposed to lethal ricin aerosols were followed clinically using telemetry, by clinical laboratory analyses and by post-mortem examination. Animals exposed to lethal aerosolized ricin developed fever associated with thermal instability, tachycardia, and dyspnea. In the peripheral blood a marked neutrophilia (without immature bands) developed at 24 h. This was accompanied by an increase in monocytes, but depletion of lymphocytes. Red cell indices indicated hemoconcentration, as did serum chemistries, with modest increases in sodium and blood urea nitrogen (BUN). Serum albumin was strikingly decreased. These observations are consistent with the pathological observations of fluid shifts to the lungs, in the form of hemorrhages, inflammatory exudates, and tissue edema. In macaques exposed to sublethal aerosols of ricin, late pathologic consequences included chronic pulmonary fibrosis, likely mediated by M2 macrophages. Early administration of supportive therapy, specific antidotes after exposure or vaccines prior to exposure have the potential to favorably alter this outcome.
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http://dx.doi.org/10.3390/toxins7062121DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4488692PMC
June 2015

Pathology of lethal and sublethal doses of aerosolized ricin in rhesus macaques.

Toxicol Pathol 2014 11;42(3):573-81. Epub 2013 Jun 11.

1Infectious Disease Aerobiology, Division of Microbiology, Tulane National Primate Research Center, Covington, Louisiana, USA.

Ricin toxin, a type 2 ribosome-inactivating protein and a category B bioterrorism agent, is produced from the seeds of castor oil plant (Ricinus communis). Chronic pathological changes in survivors of aerosolized ricin exposure have not been reported in primates. Here we compare and contrast the pathological changes manifested between rhesus macaques (RM) that succumbed to lethal dose of ricin (group I) and survivor RM exposed to low dose of ricin (group II). All animals in group I exhibited severe diffuse, necrotizing bronchiolitis and alveolitis with fibrinopurulent bronchointerstitial pneumonia, massive alveolar, perivascular and peribronchial/bronchiolar edema with hemorrhage, and necropurulent and hemorrhagic tracheobronchial lymphadenitis. All animals from group II had multifocal, fibrosing interstitial pneumonia with prominent alveolar histiocytosis and type II pneumocyte hyperplasia. Subacute changes like infiltration by lymphocytes and plasma cells and persistence of edematous fluid were occasionally present in lung and tracheobronchial lymph nodes. The changes appear to be a continuum wherein the inflammatory response shifts from an acute to subacute/chronic reparative process if the animals can survive the initial insult.
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http://dx.doi.org/10.1177/0192623313492248DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3849233PMC
December 2014

miR-375 regulates rat alveolar epithelial cell trans-differentiation by inhibiting Wnt/β-catenin pathway.

Nucleic Acids Res 2013 Apr 8;41(6):3833-44. Epub 2013 Feb 8.

Lundberg-Kienlen Lung Biology and Toxicology Laboratory, Department of Physiological Sciences, Oklahoma State University, Stillwater, OK 74078, USA.

Alveolar epithelial cell (AEC) trans-differentiation is a process where type II alveolar epithelial cells (AEC II) trans-differentiate into type I alveolar epithelial cells (AEC I) during lung recovery after various injuries, in which AEC I are damaged. This process is critical for lung tissue repair. MicroRNAs are a group of small RNAs that regulate gene expression at the post-transcriptional level. They have the potential to regulate almost every aspect of cell physiology. However, whether AEC trans-differentiation is regulated by microRNAs is completely unknown. In this study, we found that miR-375 was downregulated during AEC trans-differentiation. The overexpression of miR-375 with an adenoviral vector inhibited alveolar epithelial trans-differentiation as indicated by an increase in the AEC II marker, surfactant protein C, and decreases in the AEC I markers, T1α and advanced glycosylation end product-specific receptor. miR-375 also inhibited the Wnt/β-catenin pathway. The constitutively activation of Wnt/β-catenin signaling with a stabilized form of β-catenin blocked the miR-375 effects. Frizzled 8 was identified as a target of miR-375. In summary, our results demonstrate that miR-375 regulates AEC trans-differentiation through the Wnt/β-catenin pathway. This discovery may provide new targets for therapeutic intervention to benefit lung recovery from injuries.
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http://dx.doi.org/10.1093/nar/gks1460DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3616718PMC
April 2013

Identification of microRNAs changed in the neonatal lungs in response to hyperoxia exposure.

Physiol Genomics 2012 Oct 21;44(20):970-80. Epub 2012 Aug 21.

The Lundberg-Kienlen Lung Biology and Toxicology Laboratory, Department of Physiological Sciences, Oklahoma State University, Stillwater, Oklahoma, OK 74078, USA.

Bronchopulmonary dysplasia (BPD) is a multifactorial chronic lung disease of premature infants. BPD can be attributed to the dysregulation of normal lung development due to ventilation and oxygen toxicity, resulting in pathologic complications of impaired alveolarization and vascularization. MicroRNAs (miRNA) are small noncoding RNAs that regulate gene expression posttranscriptionally and are implicated in diverse biological processes and diseases. The objectives of this study are to identify the changed miRNAs and their target genes in neonatal rat lungs in response to hyperoxia exposure. Using miRNA microarray and real-time PCR analyses, we found downregulation of five miRNAs, miR-342, miR-335, miR-150, miR-126*, and miR-151*, and upregulation of two miRNAs, miR-21 and miR-34a. Some of these miRNAs had the highest expression during embryonic and early postnatal development. DNA microarray analysis yielded several genes with conserved binding sites for these altered miRNAs. Glycoprotein nonmetastatic melanoma protein b (GPNMB) was experimentally verified as a target of miR-150. In summary, we identified seven miRNAs that were changed in hyperoxia-exposed neonatal lungs. These results provide a basis for deciphering the mechanisms involved in the spatial and temporal regulation of proteins that contribute to the pathogenesis of BPD.
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http://dx.doi.org/10.1152/physiolgenomics.00145.2011DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3472467PMC
October 2012

What is your diagnosis? Fine-needle aspirate of a third eyelid mass in a Paint horse.

Vet Clin Pathol 2012 Jun 6;41(2):299-300. Epub 2012 Apr 6.

Department of Pathobiological Sciences, School of Veterinary Medicine, Louisiana State University, Skip Bertman Drive, Baton Rouge, LA 70803, USA.

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http://dx.doi.org/10.1111/j.1939-165X.2012.00420.xDOI Listing
June 2012

What is your diagnosis? Avian poxvirus.

J Avian Med Surg 2009 Dec;23(4):325-8

Department of Veterinary Clinical Sciences, School of Veterinary Medicine, Louisiana State University, Baton Rouge, LA 70803, USA.

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http://dx.doi.org/10.1647/1082-6742-23.4.325DOI Listing
December 2009

Pleiotrophin regulates lung epithelial cell proliferation and differentiation during fetal lung development via beta-catenin and Dlk1.

J Biol Chem 2009 Oct 6;284(41):28021-28032. Epub 2009 Aug 6.

Lundberg-Kienlen Lung Biology and Toxicology Laboratory, Department of Physiological Sciences, Oklahoma State University, Stillwater, Oklahoma 74078. Electronic address:

The role of pleiotrophin in fetal lung development was investigated. We found that pleiotrophin and its receptor, protein-tyrosine phosphatase receptor beta/zeta, were highly expressed in mesenchymal and epithelial cells of the fetal lungs, respectively. Using isolated fetal alveolar epithelial type II cells, we demonstrated that pleiotrophin promoted fetal type II cell proliferation and arrested type II cell trans-differentiation into alveolar epithelial type I cells. Pleiotrophin also increased wound healing of injured type II cell monolayer. Knockdown of pleiotrophin influenced lung branching morphogenesis in a fetal lung organ culture model. Pleiotrophin increased the tyrosine phosphorylation of beta-catenin, promoted beta-catenin translocation into the nucleus, and activated T cell factor/lymphoid enhancer factor transcription factors. Dlk1, a membrane ligand that initiates the Notch signaling pathway, was identified as a downstream target of the pleiotrophin/beta-catenin pathway by endogenous dlk1 expression, promoter assay, and chromatin immunoprecipitation. These results provide evidence that pleiotrophin regulates fetal type II cell proliferation and differentiation via integration of multiple signaling pathways including pleiotrophin, beta-catenin, and Notch pathways.
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http://dx.doi.org/10.1074/jbc.M109.052530DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC2788854PMC
October 2009

MicroRNA-127 modulates fetal lung development.

Physiol Genomics 2009 May 17;37(3):268-78. Epub 2009 Mar 17.

Lundberg-Kienlen Lung Biology and Toxicology Laboratory, Department of Physiological Sciences, Oklahoma State University, Stillwater, Oklahoma 74078, USA.

MicroRNAs (miRNAs) are small endogenous RNAs and are widely regarded as one of the most important regulators of gene expression in both plants and animals. To define the roles of miRNAs in fetal lung development, we profiled the miRNA expression pattern during lung development with a miRNA microarray. We identified 21 miRNAs that showed significant changes in expression during lung development. These miRNAs were grouped into four distinct clusters based on their expression pattern. Cluster 1 contained miRNAs whose expression increased as development progressed, while clusters 2 and 3 showed the opposite trend of expression. miRNAs in cluster 4 including miRNA-127 (miR-127) had the highest expression at the late stage of fetal lung development. Quantitative real-time PCR validated the microarray results of six selected miRNAs. In situ hybridization demonstrated that miR-127 expression gradually shifted from mesenchymal cells to epithelial cells as development progressed. Overexpression of miR-127 in fetal lung organ culture significantly decreased the terminal bud count, increased terminal and internal bud sizes, and caused unevenness in bud sizes, indicating improper development. These findings suggest that miR-127 may have an important role in fetal lung development.
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http://dx.doi.org/10.1152/physiolgenomics.90268.2008DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC2685501PMC
May 2009

Ionotropic GABA receptor expression in the lung during development.

Gene Expr Patterns 2008 Jul 3;8(6):397-403. Epub 2008 May 3.

Department of Physiological Sciences, Oklahoma State University, 264 McElroy Hall, Stillwater, OK 74078, USA.

Cl(-) transport is essential for lung development. Because gamma-aminobutyric acid (GABA) receptors allow the flow of negatively-charged Cl(-) ions across the cell membrane, we hypothesized that the expression of ionotropic GABA receptors are regulated in the lungs during development. We identified 17 GABA receptor subunits in the lungs by real-time PCR. These subunits were categorized into four groups: Group 1 had high mRNA expression during fetal stages and low in adults; Group 2 had steady expression to adult stages with a slight up-regulation at birth; Group 3 showed an increasing expression from fetal to adult lungs; and Group 4 displayed irregular mRNA fluctuations. The protein levels of selected subunits were also determined by Western blots and some subunits had protein levels that corresponded to mRNA levels. Further studied subunits were primarily localized in epithelial cells in the developing lung with differential mRNA expression between isolated cells and whole lung tissues. Our results add to the knowledge of GABA receptor expression in the lung during development.
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http://dx.doi.org/10.1016/j.gep.2008.04.008DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC2581461PMC
July 2008

Gene expression of rat alveolar type II cells during hyperoxia exposure and early recovery.

Free Radic Biol Med 2007 Aug 31;43(4):628-42. Epub 2007 May 31.

Department of Physiological Sciences, Oklahoma State University, 264 McElroy Hall, Stillwater, OK 74078, USA.

Alveolar epithelial cell (AEC) injury and repair during hyperoxia exposure and recovery have been investigated for decades, but the molecular mechanisms of these processes are not clear. To identify potentially important genes involved in lung injury and repair, we studied the gene expression profiles of isolated AEC II from control, 48-h hyperoxia-exposed (>95% O(2)), and 1-7 day recovering rats using a DNA microarray containing 10,000 genes. Fifty genes showed significant differential expression between two or more time points (P<0.05, fold change >2). These genes can be classified into 8 unique gene expression patterns. Real-time PCR verified 14 selected genes in three patterns related to hyperoxia exposure and early recovery. The change in the protein level for two of the selected genes, bmp-4 and retnla, paralleled that of the mRNA level. Many of these genes were found to be involved in cell proliferation and differentiation. In an in vitro AEC trans-differentiation culture model using AEC II isolated from control and 48-h hyperoxia-exposed rats, the expressions of the cell proliferation and differentiation genes identified above were consistent with their predicted roles in the trans-differentiation of AEC. These data indicate that a coordinated mechanism may control AEC differentiation during in vivo hyperoxia exposure and recovery as well as during in vitro AEC culture.
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http://dx.doi.org/10.1016/j.freeradbiomed.2007.05.024DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC2075096PMC
August 2007

Trans-differentiation of alveolar epithelial type II cells to type I cells involves autocrine signaling by transforming growth factor beta 1 through the Smad pathway.

J Biol Chem 2007 Feb 11;282(6):3968-76. Epub 2006 Dec 11.

Department of Physiological Sciences, Oklahoma State University, Stillwater, Oklahoma 74078, USA.

Type II alveolar epithelial cells (AEC II) proliferate and transdifferentiate into type I alveolar epithelial cells (AEC I) when the normal AEC I population is damaged in the lung alveoli. We hypothesized that signaling by transforming growth factor beta1 (TGF beta1), through its downstream Smad proteins, is involved in keeping AEC II quiescent in normal cells and its altered signaling may be involved in the trans-differentiation of AEC II to AEC I. In the normal lung, TGF beta1 and Smad4 were highly expressed in AEC II. Using an in vitro cell culture model, we demonstrated that the trans-differentiation of AEC II into AEC I-like cells began with a proliferative phase, followed by a differentiation phase. The expression of TGF beta1, Smad2, and Samd3 and their phosphorylated protein forms, and cell cycle inhibitors, p15(Ink4b) and p21(Cip1), was lower during the proliferative phase but higher during the differentiation phase. Furthermore, cyclin-dependent kinases 2, 4, and 6 showed an opposite trend of expression. TGF beta1 secretion into the media increased during the differentiation phase, indicating an autocrine regulation. The addition of TGF beta1 neutralizing antibody after the proliferative phase and silencing of Smad4 by RNA interference inhibited the trans-differentiation process. In summary, our results suggest that the trans-differentiation of AEC II to AEC I is modulated by signaling through the Smad-dependent TGF beta1 pathway by altering the expression of proteins that control the G1 to S phase entry in the cell cycle.
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http://dx.doi.org/10.1074/jbc.M609060200DOI Listing
February 2007

Alveolar type I cells protect rat lung epithelium from oxidative injury.

J Physiol 2006 May;572(Pt 3):625-38

Department of Physiological Sciences, Oklahoma State University, Stillwater, OK 74078, USA.

The lung alveolar surface is covered by two morphologically and functionally distinct cells: alveolar epithelial cell types I and II (AEC I and II). The functions of AEC II, including surfactant release, cell differentiation and ion transport, have been extensively studied. However, relatively little is known regarding the physiological functions of AEC I. Global gene expression profiling of freshly isolated AEC I and II revealed that many genes were differentially expressed in AEC I. These genes have a diversity of functions, including cell defence. Nine out of 10 selected genes were verified by quantitative real-time PCR. Two genes, apolipoprotein E (Apo E) and transferrin, were further characterized and functionally studied. Immunohistochemistry indicated that both proteins were specifically localized in AEC I. Up-regulation of Apo E and transferrin was observed in hyperoxic lungs. Functionally, Apo E and transferrin play a protective role against oxidative stress in an animal model. Our studies suggest that AEC I is not just a simple barrier for gas exchange, but a functional cell that protects alveolar epithelium from injury.
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http://dx.doi.org/10.1113/jphysiol.2005.103465DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC1779994PMC
May 2006

Expression profile of IGF system during lung injury and recovery in rats exposed to hyperoxia: a possible role of IGF-1 in alveolar epithelial cell proliferation and differentiation.

J Cell Biochem 2006 Apr;97(5):984-98

Department of Physiological Sciences, Oklahoma State University, Stillwater, Oklahoma 74078, USA.

Although several studies have shown that an induction of insulin-like growth factor (IGF) components occurs during hyperoxia-mediated lung injury, the role of these components in tissue repair is not well known. The present study aimed to elucidate the role of IGF system components in normal tissue remodeling. We used a rat model of lung injury and remodeling by exposing rats to > 95% oxygen for 48 h and allowing them to recover in room air for up to 7 days. The mRNA expression of IGF-I, IGF-II, and IGF-1 receptor (IGF-1R) increased during injury. However, the protein levels of these components remained elevated until day 3 of the recovery and were highly abundant in alveolar type II cells. Among IGF binding proteins (IGFBPs), IGFBP-5 mRNA expression increased during injury and at all the recovery time points. IGFBP-2 and -3 mRNA were also elevated during injury phase. In an in vitro model of cell differentiation, the expression of IGF-I and IGF-II increased during trans-differentiation of alveolar epithelial type II cells into type-I like cells. The addition of anti-IGF-1R and anti-IGF-I antibodies inhibited the cell proliferation and trans-differentiation to some extent, as evident by cell morphology and the expression of type I and type II cell markers. These findings demonstrate that the IGF signaling pathway plays a critical role in proliferation and differentiation of alveolar epithelium during tissue remodeling.
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http://dx.doi.org/10.1002/jcb.20653DOI Listing
April 2006

Hemoglobin is expressed in alveolar epithelial type II cells.

Biochem Biophys Res Commun 2005 Aug;333(4):1348-52

Department of Physiological Sciences, Oklahoma State University, Stillwater, OK 74078, USA.

Hemoglobin is the main oxygen carrying heme protein in erythrocytes. In an effort to study the differential gene expression of alveolar epithelial type I and type II cells using DNA microarray technique, we found that the mRNAs of hemoglobin alpha- and beta-chains were expressed in type II cells, but not in type I cells. The microarray data were confirmed by RT-PCR. The mRNA expression of both chains decreased when type II cells trans-differentiated into type I-like cells. Immunocyto/histochemistry revealed that hemoglobin protein was specifically localized in type II cells of a lung cell mixture and rat lung tissue. The endogenous synthesis of hemoglobin in alveolar epithelial cells suggests that hemoglobin may have unidentified functions other than oxygen transport in the lung.
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http://dx.doi.org/10.1016/j.bbrc.2005.06.042DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC1314978PMC
August 2005