Publications by authors named "Xiuyun Jiang"

39 Publications

Phenotypic spectrum and mechanism analysis of Schaff Yang syndrome: A case report on new mutation of MAGEL2 gene.

Medicine (Baltimore) 2021 Jun;100(24):e26309

Department of Endocrinology, Shandong Provincial Hospital affiliated to Shandong First Medical University.

Rationale: The Schaaf-Yang syndrome (SYS) is an autosomal dominant multi-system genetic disease caused by melanoma antigen L2 (MAGEL2) gene mutations imprinted by mothers and expressed by fathers on the 15q11-15q13 chromosomes in the critical region of Prader-Willi. MAGEL2 is a single exon gene and one of the protein-coding genes of the Prader-Willi domain. MAGEL2 is a matrilineal imprinted gene (i.e., the maternal chromosome is methylated). It is only expressed by unmethylated paternal alleles, and the individual is affected only when the variation occurs on the paternal allele.

Patient Concerns: We reported a patient with MAGEL2 gene new site mutation who had mild intellectual disability, social fear, small hands and feet, obesity issues, dyskinesia, growth retardation, language lag and sexual development disorder.

Diagnosis: Whole-exome sequencing showed a heterozygous variation in the MAGEL2 gene, NM_019066.4:c.1687C > T (p.Q563X) and diagnosed as Schaaf-Yang syndrome.

Interventions: Patient was advised to reduce weight, control blood lipids, blood glucose through appropriate strengthening of exercise and diet control in the future. At the same time, the family members were advised to provide mental training to the patient to strengthen the contact and communication with the outside world and improve the autistic symptoms. Because of the patient's bilateral cryptorchidism, it is recommended that the patient should be treated with bilateral cryptorchidism reduction fixation.

Outcomes: After a follow-up of the patient for 2 months, the patient is still walking unsteadily and requires an auxiliary reference material to walk normally. There is no significant change in height compared to before, and the weight has dropped by about 2 kg in the past 2 months. The symptoms of autism have improved slightly. The patient is willing to communicate with outsiders; his intelligence has not improved significantly, and his academic performance in school is still at the middle and lower levels.

Lessons: The pathogenesis of SYS is complex, involving multiple pathways such as Leptin-POMC, MAGEL2-USP7-TRIM27 complex and oxytocin. Our study has also found that certain fatal phenotypes such as respiratory distress have a high incidence at individual sites, and early detection and timely intervention may prolong the life span of patients. Therefore, for patients in whom SYS is highly suspected, gene detection should be carried out as soon as possible.
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http://dx.doi.org/10.1097/MD.0000000000026309DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8213290PMC
June 2021

Characterization of a novel COL10A1 variant associated with Schmid-type metaphyseal chondrodysplasia and a literature review.

Mol Genet Genomic Med 2021 May 25;9(5):e1668. Epub 2021 Mar 25.

Department of Endocrinology and Metabolism, Shandong Provincial Hospital, Cheeloo College of Medicine, Shandong University, Jinan, China.

Background: Schmid-type metaphyseal chondrodysplasia (SMCD) is a rare autosomal dominant skeletal dysplasia caused by heterozygous mutations in COL10A1, the gene which encodes collagen type X alpha 1 chain. However, its genotype-phenotype relationship has not been fully determined. Subjects and Methods The proband is a 2-year-old boy, born of non-consanguineous Chinese parents. We conducted a systematic analysis of the clinical and radiological characteristics and a follow-up study of the proband. Whole-exome sequencing was applied for the genetic analysis, together with bioinformatic analysis of predicted consequences of the identified variant. A homotrimer model was built to visualize the affected region and predict possible outcomes of this variant. Furthermore, a literature review and genotype-phenotype analysis were performed by online searching all cases with SMCD.

Results: A novel heterozygous variant (NM_000493.4: c.1863_1866delAATG, NP_000484.2: p.(Met622 Thrfs*54)) was identified in COL10A1 gene in the affected child. And it was predicted to be pathogenic by in silico analysis. Protein modeling revealed that the variant was located in the NC1 domain, which was predicted to produce truncated collagen and impair the trimerization of collagen type X alpha 1 chain and combination with molecules in the matrix. Moreover, genotype-phenotype correlation analysis demonstrated that patients with truncating variants or variants in NC1 domain often presented earlier onset and severer symptoms compared with those with non-truncating or variants in non-NC1 domains.

Conclusion: The NC1 domain of COL10A1 was proved to be the hotspot region underlying SMCD, patients with variants in NC1 domain were more likely to present severer manifestations at an earlier age.
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http://dx.doi.org/10.1002/mgg3.1668DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8172203PMC
May 2021

Prevalence of Nontuberculous Mycobacterial Disease in the Changchun District of China.

Curr Microbiol 2021 Apr 9;78(4):1643-1647. Epub 2021 Mar 9.

College of Animal Medicine, College of Animal Science and Technology, College of Life Science, Jilin Provincial Engineering Research Center of Animal Probiotics and Key Laboratory of Animal Production and Product Quality Safety of Ministry of Education, Jilin Agricultural University, Changchun, 130118, China.

Rates of nontuberculous mycobacterial (NTM) disease are rapidly increasing throughout the globe. NTM disease, as an emerging infectious disease, it is very important to summarize and analyze the prevalence and main pathogenic bacteria. However, there is no relevant report in Changchun district. In the present report, 8765 clinical samples were collected between January 2017 and December 2019, we reviewed patient electronic medical records and thereby summarized the causative species associated with NTM disease in the Changchun district of China. Of 8765 clinical samples, 1987 samples yielded positive cultures. Of these cultures, 1868 (94.01%) were Mycobacterium tuberculosis, 37 (1.86%) were Mycobacterium bovis, and 82 (4.13%) were NTM. A total of 84 NTM strains were isolated from these 82 cultures, with Mycobacterium intracellulare being the most prevalent isolate therein (44.05%). NTM infection status was associated with location of residence [OR (95% CI) 3.92 (1.20-12.8)]. No apparent correlations were observed between cultured NTM species and patient clinical symptoms. Bronchiectasis was the most prevalent radiographic finding associated with NTM cases [OR (95% CI) 9.00 (1.27-63.89)]. In summary, NTM disease is a growing threat to global public health, and researchers and clinicians should thus focus on the appropriate identification of NTM species and the differentiation between NTM infections and tuberculosis.
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http://dx.doi.org/10.1007/s00284-021-02422-yDOI Listing
April 2021

Coherent Electron Transport across a 3 nm Bioelectronic Junction Made of Multi-Heme Proteins.

J Phys Chem Lett 2020 Nov 3;11(22):9766-9774. Epub 2020 Nov 3.

Department of Physics and Astronomy, University College London, Gower Street, London WC1E 6BT, U.K.

Multi-heme cytochromes (MHCs) are fascinating proteins used by bacterial organisms to shuttle electrons within, between, and out of their cells. When placed in solid-state electronic junctions, MHCs support temperature-independent currents over several nanometers that are 3 orders of magnitude higher compared to other redox proteins of similar size. To gain molecular-level insight into their astonishingly high conductivities, we combine experimental photoemission spectroscopy with DFT+Σ current-voltage calculations on a representative Gold-MHC-Gold junction. We find that conduction across the dry, 3 nm long protein occurs via off-resonant coherent tunneling, mediated by a large number of protein valence-band orbitals that are strongly delocalized over heme and protein residues. This picture is profoundly different from the electron hopping mechanism induced electrochemically or photochemically under aqueous conditions. Our results imply that the current output in solid-state junctions can be even further increased in resonance, for example, by applying a gate voltage, thus allowing a quantum jump for next-generation bionanoelectronic devices.
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http://dx.doi.org/10.1021/acs.jpclett.0c02686DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7681787PMC
November 2020

Which Multi-Heme Protein Complex Transfers Electrons More Efficiently? Comparing MtrCAB from with OmcS from .

J Phys Chem Lett 2020 Nov 26;11(21):9421-9425. Epub 2020 Oct 26.

Department of Physics and Astronomy and Thomas Young Centre, University College London, London WC1E 6BT, United Kingdom.

Microbial nanowires are fascinating biological structures that allow bacteria to transport electrons over micrometers for reduction of extracellular substrates. It was recently established that the nanowires of both and are made of multi-heme proteins; but, while employs the 20-heme protein complex MtrCAB, uses a redox polymer made of the hexa-heme protein OmcS, begging the question as to which protein architecture is more efficient in terms of long-range electron transfer. Using a multiscale computational approach we find that OmcS supports electron flows about an order of magnitude higher than MtrCAB due to larger heme-heme electronic couplings and better insulation of hemes from the solvent. We show that heme side chains are an essential structural element in both protein complexes, accelerating rate-limiting electron tunnelling steps up to 1000-fold. Our results imply that the alternating stacked/T-shaped heme arrangement present in both protein complexes may be an evolutionarily convergent design principle permitting efficient electron transfer over very long distances.
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http://dx.doi.org/10.1021/acs.jpclett.0c02842DOI Listing
November 2020

An ESCRT-III Polymerization Sequence Drives Membrane Deformation and Fission.

Cell 2020 09 18;182(5):1140-1155.e18. Epub 2020 Aug 18.

Department of Biochemistry, University of Geneva, 1211 Geneva, Switzerland; National Center of Competence in Research in Chemical Biology, University of Geneva, 1211 Geneva, Switzerland. Electronic address:

The endosomal sorting complex required for transport-III (ESCRT-III) catalyzes membrane fission from within membrane necks, a process that is essential for many cellular functions, from cell division to lysosome degradation and autophagy. How it breaks membranes, though, remains unknown. Here, we characterize a sequential polymerization of ESCRT-III subunits that, driven by a recruitment cascade and by continuous subunit-turnover powered by the ATPase Vps4, induces membrane deformation and fission. During this process, the exchange of Vps24 for Did2 induces a tilt in the polymer-membrane interface, which triggers transition from flat spiral polymers to helical filament to drive the formation of membrane protrusions, and ends with the formation of a highly constricted Did2-Ist1 co-polymer that we show is competent to promote fission when bound on the inside of membrane necks. Overall, our results suggest a mechanism of stepwise changes in ESCRT-III filament structure and mechanical properties via exchange of the filament subunits to catalyze ESCRT-III activity.
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http://dx.doi.org/10.1016/j.cell.2020.07.021DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7479521PMC
September 2020

M.neoaurum infection increased the inhibitory function of Tregs and the death rate associated with Salmonella coinfection.

Res Vet Sci 2020 Oct 4;132:108-115. Epub 2020 May 4.

College of Animal Science and Technology, Jilin Provincial Engineering Research Center of Animal Probiotics and Key Laboratory of Animal Production and Product Quality Safety of Ministry of Education, Jilin Agricultural University, Changchun, China. Electronic address:

Mycobacterium neoaurum belongs to the nontuberculous mycobacteria (NTM) and is ubiquitously present in the environment. However, the changes in Treg percentages and suppressive properties in mice infected with M. neoaurum are still not elucidated. In this study, mice were intraperitoneally injected with M. neoaurum. The change in the CD4CD25 Treg cell percentage in the spleen was analyzed using flow cytometry. There was a significant increase in the number of CD4CD25 cells by week 6 postinfection, with a peak proportion of approximately 2%. The Foxp3 and IL-10 mRNA expression in CD4CD25 cells from the spleens of M.neoaurum-infected mice was higher than that in CD4CD25 cells from the spleens of noninfected controls. Proliferation suppression assay results indicated that CD4CD25 cells suppressed the proliferation of CD4CD25 cells at week 6 after M.neoaurum infection, and the suppression rate reached 89.8%. However, CD4CD25 cells from the noninfected control group did not suppress the proliferation of CD4CD25 cells. Based on the above results, mice were subjected to oral administration of S. Typhimurium at 6 weeks postinfection with M. neoaurum, and we found that the mortality of the M.neoaurum-S. Typhimurium infection group was higher than that of the S. Typhimurium infection group. In addition, serious pathological changes appeared in the liver and cecum of the M.neoaurum-S.Typhimurium infection group compared with those of the S. Typhimurium infection group. M. neoaurum increased Treg percentages and suppressed spleen function in mice. These results revealed the possibility that persistent M.neoaurum infection could increase the occurrence of secondary infection.
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http://dx.doi.org/10.1016/j.rvsc.2020.05.002DOI Listing
October 2020

Ergodicity Breaking in Thermal Biological Electron Transfer? Cytochrome C Revisited II.

J Phys Chem B 2020 04 13;124(16):3336-3342. Epub 2020 Apr 13.

Department of Physics and Astronomy and Thomas Young Centre, University College London, London WC1E 6BT, U.K.

It was recently suggested that cytochrome c operates in an ergodicity-breaking regime characterized by unusually large energy gap thermal fluctuations and associated reorganization free energies for heme oxidation of up to 3.0 eV. The large fluctuations were reported to lower activation free energy for oxidation of the heme cofactor by almost a factor of 2 compared to the case where ergodicity is maintained. Our group has recently investigated this claim computationally at several levels of theory and found no evidence for such large energy gap fluctuations. Here we address the points of our earlier work that have raised criticism and we also extend our previous investigation by considering a simple linear polarizability model for cytochrome c oxidation. We find very consistent results among all our computational approaches, ranging from classical molecular dynamics, to the linear polarizability model to QM(PMM)/MM to full QM(DFT)/MM electrostatic emdedding. None of them support the notion of very large energy gap fluctuations or ergodicity breaking. The deviation between our simulations and the ones reported in [ 2017, 121, 4958] is traced back to rather large electric fields at the Fe site of the heme c cofactor in that study, not seen in our simulations, neither with the AMBER nor with the CHARMM force field. While ergodicity breaking effects may well occur in other biological ET, our numerical evidence suggests that this is not the case for cytochrome c.
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http://dx.doi.org/10.1021/acs.jpcb.0c01414DOI Listing
April 2020

A Phase II Trial of Pembrolizumab and Vorinostat in Recurrent Metastatic Head and Neck Squamous Cell Carcinomas and Salivary Gland Cancer.

Clin Cancer Res 2020 02 3;26(4):837-845. Epub 2019 Dec 3.

Division of Oncology, Department of Medicine, University of Washington, Seattle, Washington.

Purpose: This clinical trial combined pembrolizumab and vorinostat in recurrent/metastatic squamous cell carcinomas of the head and neck (HN), and salivary gland cancer (SGC).

Patients And Methods: Patients with progressing incurable HN and SGC, Eastern Cooperative Oncology Group (ECOG) ≤1, no prior immunotherapy, RECIST1.1 measurable disease, and normal organ function were eligible. Pembrolizumab 200 mg was given intravenous every 21 days, and vorinostat 400 mg given orally 5 days on and 2 days off during each 21-day cycle. Primary endpoints were safety and objective response rates.

Results: From November 2015 to August 2017, 25 patients with HN and 25 SGC were enrolled. Median age was 61 (range, 33-86) years, 39 (78%) were male, 21 (62%) were never smokers, and 27 (54%) had ECOG 0. In HN, 13 (52%) were p16+ oropharynx. Most common SGC histologies were adenoid cystic 12 (48%), acinic cell 3 (12%), and mucoepidermoid 3 (12%). Adverse events (AEs) in all patients were: 27 (54%) with grade ≥ 1 and 18 (36%) with grade ≥ 3. The most common AEs in all patients were renal insufficiency in seven, (14%), fatigue in six, (12%), and nausea in three (6%). There were three (12%) deaths on study. Responses in HN were complete response (CR) 0, partial response (PR) eight (32%), and stable disease (SD) five (20%). Efficacy in SGCs was CR 0, PR four (16%) in one lymphoepithelioma-like carcinoma, two acinic cell, one adenoid cystic, and SD 14 (56%). In the HN group, median follow-up (mFUP) was 12.6 months, median overall survival (mOS) was 12.6 months, and median progression-free survival (mPFS) was 4.5 months. In SGC, mFUP was 13.1 months, mOS was 14.0 months, and mPFS was 6.9 months.

Conclusions: This combination demonstrated activity in HN, with fewer responses in SGC. Toxicities were higher than reported with pembrolizumab alone.
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http://dx.doi.org/10.1158/1078-0432.CCR-19-2214DOI Listing
February 2020

Ultrafast Light-Driven Electron Transfer in a Ru(II)tris(bipyridine)-Labeled Multiheme Cytochrome.

J Am Chem Soc 2019 09 10;141(38):15190-15200. Epub 2019 Sep 10.

School of Chemistry and School of Biological Sciences , University of East Anglia , Norwich Research Park , Norwich NR4 7TJ , United Kingdom.

Multiheme cytochromes attract much attention for their electron transport properties. These proteins conduct electrons across bacterial cell walls and along extracellular filaments and when purified can serve as bionanoelectronic junctions. Thus, it is important and necessary to identify and understand the factors governing electron transfer in this family of proteins. To this end we have used ultrafast transient absorbance spectroscopy, to define heme-heme electron transfer dynamics in the representative multiheme cytochrome STC from in aqueous solution. STC was photosensitized by site-selective labeling with a Ru(II)(bipyridine) dye and the dynamics of light-driven electron transfer described by a kinetic model corroborated by molecular dynamics simulation and density functional theory calculations. With the dye attached adjacent to STC Heme IV, a rate constant of 87 × 10 s was resolved for Heme IV → Heme III electron transfer. With the dye attached adjacent to STC Heme I, at the opposite terminus of the tetraheme chain, a rate constant of 125 × 10 s was defined for Heme I → Heme II electron transfer. These rates are an order of magnitude faster than previously computed values for unlabeled STC. The Heme III/IV and I/II pairs exemplify the T-shaped heme packing arrangement, prevalent in multiheme cytochromes, whereby the adjacent porphyrin rings lie at 90° with edge-edge (Fe-Fe) distances of ∼6 (11) Å. The results are significant in demonstrating the opportunities for pump-probe spectroscopies to resolve interheme electron transfer in Ru-labeled multiheme cytochromes.
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http://dx.doi.org/10.1021/jacs.9b06858DOI Listing
September 2019

Ergodicity-Breaking in Thermal Biological Electron Transfer? Cytochrome C Revisited.

J Phys Chem B 2019 09 26;123(35):7588-7598. Epub 2019 Aug 26.

Department of Physics and Astronomy and Thomas Young Centre, University College London, London WC1E 6BT, United Kingdom.

It was recently suggested that certain redox proteins operate in an ergodicity-breaking regime to facilitate biological electron transfer (ET). A signature for this is a large variance reorganization free energy (several electronvolts) but a significantly smaller Stokes reorganization free energy due to incomplete protein relaxation on the time scale of the ET event. Here we investigate whether this picture holds for oxidation of cytochrome c in aqueous solution, at various levels of theory including classical molecular dynamics with two additive and one electronically polarizable force field, and QM/MM calculations with the QM region treated by full electrostatic DFT embedding and by the perturbed matrix method. Sampling the protein and energy gap dynamics over more than 250 ns, we find no evidence for ergodicity-breaking effects. In particular, the inclusion of electronic polarizability of the heme group at QM/MM levels did not induce nonergodic effects, contrary to previous reports by Matyushov et al. The well-known problem of overestimation of reorganization free energies with additive force fields is cured when the protein and solvent are treated as electronically polarizable. Ergodicity-breaking effects may occur in other redox proteins, and our results suggest that long simulations, ideally on the ET time scale, with electronically polarizable force fields are required to obtain strong numerical evidence for them.
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http://dx.doi.org/10.1021/acs.jpcb.9b05253DOI Listing
September 2019

Mobilization of CD8 T Cells via CXCR4 Blockade Facilitates PD-1 Checkpoint Therapy in Human Pancreatic Cancer.

Clin Cancer Res 2019 Jul 2;25(13):3934-3945. Epub 2019 Apr 2.

Department of Surgery, University of Washington, Seattle, Washington.

Purpose: Pancreatic ductal adenocarcinoma (PDA) is rarely cured, and single-agent immune checkpoint inhibition has not demonstrated clinical benefit despite the presence of large numbers of CD8 T cells. We hypothesized that tumor-infiltrating CD8 T cells harbor latent antitumor activity that can be reactivated using combination immunotherapy.

Experimental Design: Preserved human PDA specimens were analyzed using multiplex IHC (mIHC) and T-cell receptor (TCR) sequencing. Fresh tumor was treated in organotypic slice culture to test the effects of combination PD-1 and CXCR4 blockade. Slices were analyzed using IHC, flow cytometry, and live fluorescent microscopy to assess tumor kill, in addition to T-cell expansion and mobilization.

Results: mIHC demonstrated fewer CD8 T cells in juxtatumoral stroma containing carcinoma cells than in stroma devoid of them. Using TCR sequencing, we found clonal expansion in each tumor; high-frequency clones had multiple DNA rearrangements coding for the same amino acid binding sequence, which suggests response to common tumor antigens. Treatment of fresh human PDA slices with combination PD-1 and CXCR4 blockade led to increased tumor cell death concomitant with lymphocyte expansion. Live microscopy after combination therapy demonstrated CD8 T-cell migration into the juxtatumoral compartment and rapid increase in tumor cell apoptosis.

Conclusions: Endogenous tumor-reactive T cells are present within the human PDA tumor microenvironment and can be reactivated by combined blockade of PD-1 and CXCR4. This provides a new basis for the rational selection of combination immunotherapy for PDA..
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http://dx.doi.org/10.1158/1078-0432.CCR-19-0081DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6606359PMC
July 2019

Kinetics of trifurcated electron flow in the decaheme bacterial proteins MtrC and MtrF.

Proc Natl Acad Sci U S A 2019 02 12;116(9):3425-3430. Epub 2019 Feb 12.

Department of Physics and Astronomy, University College London, London WC1E 6BT, United Kingdom;

The bacterium has evolved a sophisticated electron transfer (ET) machinery to export electrons from the cytosol to extracellular space during extracellular respiration. At the heart of this process are decaheme proteins of the Mtr pathway, MtrC and MtrF, located at the external face of the outer bacterial membrane. Crystal structures have revealed that these proteins bind 10 c-type hemes arranged in the peculiar shape of a staggered cross that trifurcates the electron flow, presumably to reduce extracellular substrates while directing electrons to neighboring multiheme cytochromes at either side along the membrane. Especially intriguing is the design of the heme junctions trifurcating the electron flow: they are made of coplanar and T-shaped heme pair motifs with relatively large and seemingly unfavorable tunneling distances. Here, we use electronic structure calculations and molecular simulations to show that the side chains of the heme rings, in particular the cysteine linkages inserting in the space between coplanar and T-shaped heme pairs, strongly enhance electronic coupling in these two motifs. This results in an [Formula: see text]-fold speedup of ET steps at heme junctions that would otherwise be rate limiting. The predicted maximum electron flux through the solvated proteins is remarkably similar for all possible flow directions, suggesting that MtrC and MtrF shuttle electrons with similar efficiency and reversibly in directions parallel and orthogonal to the outer membrane. No major differences in the ET properties of MtrC and MtrF are found, implying that the different expression levels of the two proteins during extracellular respiration are not related to redox function.
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http://dx.doi.org/10.1073/pnas.1818003116DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6397555PMC
February 2019

Clinical Diagnosis of X-Linked Spondyloepiphyseal Dysplasia Tarda and a Novel Missense Mutation in the Sedlin Gene (SEDL).

Int J Endocrinol 2018 10;2018:8263136. Epub 2018 Dec 10.

Department of Endocrinology, Shandong Provincial Hospital affiliated to Shandong University, China.

Objective: Spondyloepiphyseal dysplasia tarda (SEDT) is a rare hereditary bone disease characterized by spinal and epiphyseal anomalies. We identified the disease by gene sequencing in a Chinese pedigree with SEDT.

Methods: We extracted genomic DNA from five members of a four-generation Chinese SEDT kindred with three affected males and then analyzed the genetic mutation by PCR and DNA sequencing.

Results: DNA sequencing showed that the genetic missense mutation occurred one bp upstream of exon 6 in the gene in two families, and a heterozygous mutation was found in a female carrier. In addition, no mutation was found in the other members of the family.

Conclusion: SEDT in this family was caused by a G/C missense mutation in exon 6 of the gene, previously not shown to be associated with X-linked SEDT.
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http://dx.doi.org/10.1155/2018/8263136DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6311833PMC
December 2018

Establishment of Slice Cultures as a Tool to Study the Cancer Immune Microenvironment.

Methods Mol Biol 2019 ;1884:283-295

Department of Surgery, University of Washington School of Medicine, Seattle, WA, USA.

Although immunotherapy is currently being widely applied to treat a variety of cancers, there is great heterogeneity in the response to these treatments. Many in the field hypothesize that this may be attributable to the characteristics of each individual tumor immune microenvironment, in addition to systemic immune factors. Therefore, understanding the immune cell microenvironment in a variety of tumors is critically important. Specifically, the interactions among immune, stromal, and cancer cells, along with other factors in tumors, may hold the key to developing rational personalized combinations of immunotherapeutic drugs. We recently developed an organotypic slice culture technique, which enables precise study of the pancreatic ductal adenocarcinoma (PDA) tumor microenvironment. We used a Vibratome to cut fresh human tumor tissue into 250 μm thick slices, and cultured slices on cell culture inserts with 0.4 μm pore to produce our tumor slice culture (TSC) system. We showed that TSC maintained many elements of the original tumor microenvironment and architecture for approximately one week. Using this slice culture technique for PDA, we demonstrated that immune cells, including T cells and macrophages, cancer cells, and stromal myofibroblasts were present throughout the culture period. TSCs were functionally responsive to drug treatment. Live PDA slices could be stained for multicolor immunofluorescence imaging of each of the primary cellular constituents of the tumor. Finally, autologous CFSE-labeled splenocytes were observed to readily migrate into cocultured tumor slices.
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http://dx.doi.org/10.1007/978-1-4939-8885-3_20DOI Listing
June 2019

Direct evidence for heme-assisted solid-state electronic conduction in multi-heme -type cytochromes.

Chem Sci 2018 Oct 27;9(37):7304-7310. Epub 2018 Jul 27.

Department of Materials and Interfaces , Weizmann Institute of Science , Rehovot , Israel . Email:

Multi-heme cytochrome c (Cyt) proteins are key for transferring electrons out of cells, to enable intracellular oxidation to proceed in the absence of O. In these proteins most of the hemes are arranged in a linear array suggesting a facile path for electronic conduction. To test this, we studied solvent-free electron transport across two multi-heme Cyt-type proteins: MtrF (deca-heme Cyt) and STC (tetra-heme Cyt). Transport is measured across monolayers of these proteins in a solid state configuration between Au electrodes. Both proteins showed 1000× higher conductance than single heme, or heme-free proteins, but similar conductance to monolayers of conjugated organics. Conductance is found to be temperature-independent (320-80 K), suggesting tunneling as the transport mechanism. This mechanism is consistent with - curves modelling, results of which could be interpreted by having protein-electrode coupling as rate limiting, rather than transport within the proteins.
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http://dx.doi.org/10.1039/c8sc01716fDOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6166575PMC
October 2018

MiR-1180-5p regulates apoptosis of Wilms' tumor by targeting .

Onco Targets Ther 2018 16;11:823-831. Epub 2018 Feb 16.

Department of Internal Medicine, The People's Hospital of Zhoukou, Zhoukou, People's Republic of China.

Introduction: Wilms' tumor (WT), the most common childhood tumor, occurs in sporadic or familial forms. Recent findings reported that abnormal expression in microRNA (miRNA) suggests an important role of miRNAs during WT progress. MiRNAs are endogenous short-chain noncoding RNAs, which have been reported as key biomarkers for detecting tumor onset and progression. However, the functional role of miR-1180 in WT has remained unknown.

Materials And Methods: MTT and clonogenic survival assays were used to detect WT cell proliferation. Flow cytometry Annexin V-FITC was used to measure apoptosis. In addition, proteins expressions in the cells were determined by Western blotting.

Results: In the present study, we demonstrated that miR-1180 is upregulated in WT when compared with adjacent tissues by quantitative reverse-transcription polymerase chain reaction. In addition, the inhibition of miR-1180 induced apoptosis in SK-NEP-1 cell line in vitro. Moreover, luciferase reporter assay showed that protein was the target of miR-1180, which was confirmed by the results of Western blotting. Finally, in vivo data indicated that the tumor growth in mice was significantly inhibited by miR-1180 inhibitor.

Conclusion: Our results indicate that miR-1180 might serve as a therapeutic target for future WT therapy.
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http://dx.doi.org/10.2147/OTT.S148684DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5820468PMC
February 2018

Cysteine Linkages Accelerate Electron Flow through Tetra-Heme Protein STC.

J Am Chem Soc 2017 12 17;139(48):17237-17240. Epub 2017 Nov 17.

Department of Physics and Astronomy, University College London , London WC1E 6BT, U.K.

Multi-heme proteins have attracted much attention recently due to their prominent role in mediating extracellular electron transport (ET), but one of their key fundamental properties, the rate constants for ET between the constituent heme groups, have so far evaded experimental determination. Here we report the set of heme-heme theoretical ET rate constants that define electron flow in the tetra-heme protein STC by combining a novel projector-operator diabatization approach for electronic coupling calculation with molecular dynamics simulation of ET free energies. On the basis of our calculations, we find that the protein limited electron flux through STC in the thermodynamic downhill direction (heme 1→4) is ∼3 × 10 s. We find that cysteine linkages inserting in the space between the two terminal heme pairs 1-2 and 3-4 significantly enhance the overall electron flow, by a factor of about 37, due to weak mixing of the sulfur 3p orbital with the Fe-heme d orbitals. While the packing density model, and to a higher degree, the pathway model of biological ET partly capture the predicted rate enhancements, our study highlights the importance of the atomistic and chemical nature of the tunneling medium at short biological tunneling distances. Cysteine linkages are likely to enhance electron flow also in the larger deca-heme proteins MtrC and MtrF, where heme-heme motifs with sub-optimal edge-to-edge distances are used to shuttle electrons in multiple directions.
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http://dx.doi.org/10.1021/jacs.7b08831DOI Listing
December 2017

Long-lived pancreatic ductal adenocarcinoma slice cultures enable precise study of the immune microenvironment.

Oncoimmunology 2017;6(7):e1333210. Epub 2017 May 25.

Department of Surgery, University of Washington School of Medicine, Seattle, WA, USA.

Pancreatic ductal adenocarcinoma (PDA) remains a deadly disease that is rarely cured, despite many recent successes with immunotherapy for other malignancies. As the human disease is heavily infiltrated by effector T cells, we postulated that accurately modeling the PDA immune microenvironment would allow us to study mechanisms of immunosuppression that could be overcome for therapeutic benefit. Using viable precision-cut slices from fresh PDA, we developed an organotypic culture system for this purpose. We confirmed that cultured slices maintain their baseline morphology, surface area, and microenvironment after at least 6 d in culture, and demonstrated slice survival by MTT assay and by immunohistochemistry staining with Ki-67 and cleaved-Caspase-3 antibodies. Immune cells, including T cells (CD3, CD8, and FOXP3) and macrophages (CD68, CD163 and HLA-DR), as well as stromal myofibroblasts (αSMA) were present throughout the culture period. Global profiling of the PDA proteome before and after 6 d slice culture indicated that the majority of the immunological proteins identified remain stable during the culture process. Cytotoxic effects of drug treatment (staurosporine, STS and cycloheximide, CHX) on PDA slices culture confirmed that this system can be used to assess functional response and cell survival following drug treatment in both a treatment time- and dose-dependent manner. Using multicolor immunofluorescence, we stained live slices for both cancer cells (EpCAM) and immune cells (CD11b and CD8). Finally, we confirmed that autologous CFSE-labeled splenocytes readily migrate into co-cultured tumor slices. Thus, our present study demonstrates the potential to use tumor slice cultures to study the immune microenvironment of PDA.
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http://dx.doi.org/10.1080/2162402X.2017.1333210DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5543820PMC
May 2017

Biopolymers codelivering engineered T cells and STING agonists can eliminate heterogeneous tumors.

J Clin Invest 2017 Jun 24;127(6):2176-2191. Epub 2017 Apr 24.

Clinical Research Division, Fred Hutchinson Cancer Research Center, Seattle, Washington, USA.

Therapies using T cells that are programmed to express chimeric antigen receptors (CAR T cells) consistently produce positive results in patients with hematologic malignancies. However, CAR T cell treatments are less effective in solid tumors for several reasons. First, lymphocytes do not efficiently target CAR T cells; second, solid tumors create an immunosuppressive microenvironment that inactivates T cell responses; and third, solid cancers are typified by phenotypic diversity and thus include cells that do not express proteins targeted by the engineered receptors, enabling the formation of escape variants that elude CAR T cell targeting. Here, we have tested implantable biopolymer devices that deliver CAR T cells directly to the surfaces of solid tumors, thereby exposing them to high concentrations of immune cells for a substantial time period. In immunocompetent orthotopic mouse models of pancreatic cancer and melanoma, we found that CAR T cells can migrate from biopolymer scaffolds and eradicate tumors more effectively than does systemic delivery of the same cells. We have also demonstrated that codelivery of stimulator of IFN genes (STING) agonists stimulates immune responses to eliminate tumor cells that are not recognized by the adoptively transferred lymphocytes. Thus, these devices may improve the effectiveness of CAR T cell therapy in solid tumors and help protect against the emergence of escape variants.
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http://dx.doi.org/10.1172/JCI87624DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5451231PMC
June 2017

Expression and Enzyme Activity Detection of a Sepiapterin Reductase Gene from Musca domestica Larva.

Appl Biochem Biotechnol 2017 Feb 4;181(2):604-612. Epub 2016 Oct 4.

College of Animal Science and Technology, Jilin Agricultural University, Xincheng Street No.2888, Changchun, 130118, China.

Tetrahydrobiopterin (BH4) is an essential cofactor for aromatic acid hydroxylases and nitric oxide synthase. Sepiapterin reductase (SPR) catalyzes the final steps of BH4 biosynthesis. Studies on SPR from several insects and other organisms have been reported. However, thus far, enzyme activity of SPR in Musca domestica is kept unknown. In this study, 186 differentially expressed genes including SPR gene from Musca domestica (MDSPR) were screened in subtractive cDNA library. The MDSPR gene was cloned, and the recombinant MDSPI16 protein was expressed as a 51-kDa protein in soluble form. The MDSPR exhibited strong activity to the substrate sepiapterin (SP). The values of V and K of the MDSPR for SP were 6.83 μM/min and 23.48 μM, and the optimum temperature and pH of MDSPR were 50 °C and 4.0, respectively. This study provides new hypotheses and methods for the production of BH4 using insect-derived SPR.
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http://dx.doi.org/10.1007/s12010-016-2235-0DOI Listing
February 2017

A serine protease inhibitor from Musca domestica larva exhibits inhibitory activity against elastase and chymotrypsin.

Biotechnol Lett 2016 Jul 4;38(7):1147-53. Epub 2016 Apr 4.

College of Animal Science and Technology, Jilin Agricultural University, Xincheng Street No. 2888, Changchun, 130118, China.

Objective: Insect-derived serine protease inhibitors (serpins) exhibit multiple inhibitory activities, but so far, no functional roles for serpins of Musca domestica have been identified. Here, the functional features of M. domestica serine protease inhibitor (MDSPI16) were characterized.

Results: Hundred forty seven differentially expressed genes including the MDSPI16 gene were screened by constructing the subtractive cDNA library. The 1154-bp full-length MDSPI16 gene was cloned, and the recombinant MDSPI16 serpin protein was expressed as a 42.6 kDa protein in an Escherichia coli expression system. The recombinant MDSPI16 protein was purified using Ni-NTA affinity chromatography, and the inhibitory activity of MDSPI16 was assessed. MDSPI16 did not inhibit trypsin, papain, or proteinase K but strongly inhibited elastase (Ki = 2.8 nM) and chymotrypsin (Ki = 28 nM). The inhibitory activity of MDSPI16 remained stable over from 37 to 100 °C and from pH 2 to 12.

Conclusions: The MDSPI16 exhibited inhibitory activity against elastase and chymotrypsin and the inhibitory activity remained stable.
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http://dx.doi.org/10.1007/s10529-016-2089-0DOI Listing
July 2016

Identification of two novel mutations in SLC12A3 gene in two Chinese pedigrees with Gitelman syndrome and review of literature.

Clin Endocrinol (Oxf) 2015 Dec 15;83(6):985-93. Epub 2015 Jun 15.

Department of Endocrinology, Shandong Provincial Hospital affiliated to Shandong University, Jinan, Shandong, China.

Objective: Gitelman syndrome (GS) is one of the most common causes of inherited hypokalaemia. As it was caused by mutations in the SLC12A3 gene, GS is a highly heterogeneous disease. Here, we aimed to investigate the clinical and genetic characteristics of two Chinese pedigrees and summarize the advance in GS genetics, diagnosis and management.

Subjects And Methods: Two three-generation families with GS were identified and screened for mutations in the SLC12A3 gene. Genotype-phenotype correlations were analysed.

Results: The two probands (A and B) were characterized by hypokalaemia, hypomagnesaemia and hypocalciuria without hypertension. Complete DNA sequencing of the SLC12A3 gene revealed two novel compound heterozygous mutations (c.179C>T and c.234delG; c.486-490delTACGGinsA and c.1925G>A), which are predicted to drastically affect normal protein structure. The female members of the pedigrees showed mild-to-no phenotype, although they carried the same mutations as the probands. Moreover, proband B presented with more severe symptoms than did proband A, which might be related to a lower serum magnesium level. During the 1-year follow-up, both probands showed satisfactory symptom improvement following the use of potassium and magnesium supplements.

Conclusion: Our findings strongly suggested that the two novel mutations in the SLC12A3 gene are the causative agents of GS, which may provide further insights into the function of this gene and help clinicians better understand this disorder.
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http://dx.doi.org/10.1111/cen.12820DOI Listing
December 2015

Thyroid-stimulating hormone decreases HMG-CoA reductase phosphorylation via AMP-activated protein kinase in the liver.

J Lipid Res 2015 May 23;56(5):963-71. Epub 2015 Feb 23.

Departments of Endocrinology, Shandong Provincial Hospital Affiliated to Shandong University, Jinan 250021, China Institute of Endocrinology and Metabolism, Shandong Academy of Clinical Medicine, Shandong Provincial Hospital Affiliated to Shandong University, Jinan 250021, China.

Cholesterol homeostasis is strictly regulated through the modulation of HMG-CoA reductase (HMGCR), the rate-limiting enzyme of cholesterol synthesis. Phosphorylation of HMGCR inactivates it and dephosphorylation activates it. AMP-activated protein kinase (AMPK) is the major kinase phosphorylating the enzyme. Our previous study found that thyroid-stimulating hormone (TSH) increased the hepatocytic HMGCR expression, but it was still unclear whether TSH affected hepatic HMGCR phosphorylation associated with AMPK. We used bovine TSH (bTSH) to treat the primary mouse hepatocytes and HepG2 cells with or without constitutively active (CA)-AMPK plasmid or protein kinase A inhibitor (H89), and set up the TSH receptor (Tshr)-KO mouse models. The p-HMGCR, p-AMPK, and related molecular expression were tested. The ratios of p-HMGCR/HMGCR and p-AMPK/AMPK decreased in the hepatocytes in a dose-dependent manner following bTSH stimulation. The changes above were inversed when the cells were treated with CA-AMPK plasmid or H89. In Tshr-KO mice, the ratios of liver p-HMGCR/HMGCR and p-AMPK/AMPK were increased relative to the littermate wild-type mice. Consistently, the phosphorylation of acetyl-CoA carboxylase, a downstream target molecule of AMPK, increased. All results suggested that TSH could regulate the phosphorylation of HMGCR via AMPK, which established a potential mechanism for hypercholesterolemia involved in a direct action of the TSH in the liver.
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http://dx.doi.org/10.1194/jlr.M047654DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4409286PMC
May 2015

Branched-chain amino acids supplementation protects streptozotocin-induced insulin secretion and the correlated mechanism.

Biofactors 2015 Mar-Apr;41(2):127-33. Epub 2014 Oct 30.

Department of Endocrinology, Shandong Provincial Hospital Affiliated to Shandong University, Jinan, China.

Significant evidence demonstrates that oxidative stress can impair insulin secretion and contribute to the development of type 2 diabetes. Branched-chain amino acids (BCAAs) are reported to be positively related to insulin secretion. This study aimed to determine how oxidative stress affects the function of islets and whether BCAAs can ameliorate the oxidative stress, and accompanying c-jun N-terminal kinase (JNK), protein kinase D1 (PKD1), and pancreatic/duodenal homeobox-1 (PDX-1) changes induced by streptozotocin (STZ). Plasma glucose, plasma insulin, and JNK, PKD1 and PDX-1 mRNA and protein expression were measured in rats treated with STZ and BCAAs. The glucose level in STZ-induced diabetic rats was much higher than that in control animals, and the elevated plasma glucose level in diabetic rats could be significantly inhibited by BCAAs treatment. Consistent with the change in glucose levels, the levels of insulin were also affected by BCAAs treatment. The mRNA and protein expression of JNK, PDX-1, and PKD1 were significantly altered in diabetic rats compared with the control group (P<0.01) and treatment with a low dose of BCAA reversed these changes in those above markers significantly (P<0.01). The present study demonstrated that STZ-induced oxidative stress could reduce serum insulin levels and alter the JNK, PDX-1, and PKD1 expression. BCAAs restored the levels of serum insulin reversed changes in JNK, PDX-1, and PKD1 expression.
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http://dx.doi.org/10.1002/biof.1188DOI Listing
January 2016

Chronic leucine exposure results in reduced but reversible glucose-stimulated insulin secretion in INS-1 cells.

Mol Med Rep 2014 Jun 8;9(6):2554-8. Epub 2014 Apr 8.

Department of Endocrinology, Shandong Provincial Hospital Affiliated to Shandong University, Jinan, Shandong 250021, P.R. China.

Previous studies have demonstrated that sustained high leucine exposure decreases glucose-stimulated insulin secretion (GSIS). However, whether this effect is recoverable following the removal of leucine is unclear. Pancreatic/duodenal homeobox-1 (PDX-1) and its downstream target, glucose transporter 2 (GLUT2), are reported to be positively associated with insulin secretion. However, it also remains unclear whether the effect of leucine on GSIS is accompanied by alterations in PDX-1 and GLUT2. In the present study, insulin secretion, insulin content, PDX-1 and GLUT2 protein expression in INS-1 (rat insulinoma cell line) cells were assessed following a 24-h incubation in 40 mmol/l leucine. Half of the cells were incubated in leucine-free media for a further 24 h to observe the abovementioned effects. In contrast to the control, 40 mmol/l leucine for 24 or 48 h diminished GSIS at high glucose concentrations by 11% (P=0.026) or 22% (P=0.003), insulin content by 14% (P=0.008) or 20% (P=0.002), as well as decreasing PDX-1 and GLUT2 expression. When leucine was removed from the media for a further 24-h incubation, in comparison with those cells that were maintained in leucine treatment for 24 and 48 h, the high GSIS increased by 13% (P=0.032) and 27% (P=0.002), insulin content was augmented by 10% (P=0.014) and 20% (P=0.003), and the protein expression of PDX-1 and GLUT2 also increased. The present study demonstrates that sustained high concentrations of leucine induce a reversible impairment of GSIS and alter insulin content, which is mediated by PDX-1 and GLUT2, in INS-1 cells.
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http://dx.doi.org/10.3892/mmr.2014.2122DOI Listing
June 2014

Expression profiles of six circulating microRNAs critical to atherosclerosis in patients with subclinical hypothyroidism: a clinical study.

J Clin Endocrinol Metab 2014 May 11;99(5):E766-74. Epub 2014 Feb 11.

Departments of Endocrinology (X.Z., S.S., H.G., C.Y., X.J., L.G., J.Z.), Sonography (Y.Y.), and Clinic Laboratory (Z.L.), and Central Laboratory (L.G.), Shandong Provincial Hospital Affiliated to Shandong University, and Shandong Clinical Medical Center of Endocrinology and Metabolism (X.Z., S.S., H.G., C.Y., X.J., J.Z.), Institute of Endocrinology and Metabolism, Shandong Academy of Clinical Medicine, Jinan 250021, China; Department of Endocrinology (X.Z., Y.D.), the Affiliated Hospital of Taishan Medical University, Tai'an 271000, China; Department of Endocrinology (H.G.), Affiliated Hospital of Jining Medical University, Jining 272029, China; and Department of Preventive Medicine (C.W.), Shandong University of Traditional Chinese Medicine, Jinan 250355, China.

Context: Increasing evidence shows that subclinical hypothyroidism (SCH) is associated with atherosclerosis (ATH), but the association remains controversial. MicroRNAs (miRNAs) have been proved to be involved in atherosclerosis and dyslipidemia as gene regulators.

Objective: The objective of the study was to determine the expression profiles of six serum miRNAs critical to atherosclerosis in SCH patients and reanalyze the association between atherosclerosis and SCH from a new perspective. OUTCOMES, DESIGN, AND PARTICIPANTS: MicroRNA profiling analysis was performed by real-time PCR in normal control subjects (NC; n = 22); patients with subclinical hypothyroidism alone (SCH; n = 20); SCH patients plus atherosclerosis (SCH+ATH; n = 21); and patients with atherosclerosis but without subclinical hypothyroidism (ATH; n = 22).

Results: MiR-21-5p was up-regulated in SCH, SCH+ATH, and ATH groups than in the NC group. In addition, expression levels of miR-21-5p in SCH+ATH group were higher than in SCH alone and ATH alone groups, respectively. Both miR-125a-5p and miR-126-3p showed a decreased trend from NC to SCH and then to SCH+ATH or ATH subjects. MiR-221-3p and miR-222-3p were decreased in the SCH+ATH and ATH groups compared with either the NC or SCH groups. No differences were found in the levels of miR125a-5p, miR126-3p, miR221-3p, and miR222-3p between the ATH and SCH+ATH group.

Conclusions: MiR-21-5p showed the most specific expression patterns in all patients with subclinical hypothyroidism (SCH and SCH+ATH groups). Down-regulation of miR-125a-5p, miR-126-3p, miR-221-3p, and miR-222-3p may be a manifestation of atherosclerosis either in SCH+ATH or in ATH-alone patients. MiR-126-3p has the most specific expression patterns in all atherosclerosis patients (SCH+ATH and ATH groups).
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http://dx.doi.org/10.1210/jc.2013-1629DOI Listing
May 2014

Association between thyroid hormones and body fat in euthyroid subjects.

Clin Endocrinol (Oxf) 2014 Apr 23;80(4):585-90. Epub 2013 Sep 23.

Department of Endocrinology, Provincial Hospital Affiliated to Shandong University, Jinan, China; Department of Endocrinology, Yantai Yuhuangding Hospital Affiliated to Qingdao University, Yantai, China.

Objective: Thyroid hormones disorders are associated with changes of body composition. However, the relationship between thyroid hormones and body fat in a euthyroid population is unclear. The aim of this study was to explore the association between thyroid hormones and body fat in a euthyriod population.

Subjects: A total of 865 euthyroid individuals were recruited in this study. Subjects with thyroid diseases or diabetes and who were taking medications that could influence thyroid hormones, weight or glucose metabolism were excluded.

Measurements: FT3, FT4, lipid parameters, fasting insulin (FINS) and blood glucose were determined, and homeostasis model assessment for insulin resistance (HOMA-IR) was calculated. Anthropometric measurements were taken, and body fat parameters were assessed.

Results: Serum FT3 was slightly higher in body mass index (BMI) ≥25 kg/m(2) group than that in BMI <25 kg/m(2) group (P < 0·05). However, the difference was too small to have clinical significance. FT4 levels were not significantly different between the two groups. Body weight, BMI, waist circumference, hip circumference, waist-to-hip ratio (WHR), percentage of body fat (PBF), waist-fat-to-hip-fat ratio, FINS and HOMA-IR increased linearly with the elevation of FT3 adjusted for age and gender. A multivariate linear regression analysis revealed that fat mass, PBF, HOMA-IR and FT4 contributed significantly to FT3 levels.

Conclusion: Grouped according to BMI, overweight and obese subjects have similar thyroid hormones compared to those with normal weight. Body composition parameters increase with the elevation of FT3, and FT3 is associated with body fat parameters in euthyroid subjects.
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http://dx.doi.org/10.1111/cen.12311DOI Listing
April 2014

Decreased fasting blood glucose is associated with impaired hepatic glucose production in thyroid-stimulating hormone receptor knockout mice.

Endocr J 2013 10;60(8):941-50. Epub 2013 May 10.

Department of Endocrinology, Provincial Hospital Affiliated to Shandong University, Jinan, China.

Our previous study reported that thyroid-stimulating hormone (TSH) promotes cholesterol synthesis via the cyclic adenosine monophosphate/protein kinase A/cAMP regulatory element-binding protein (cAMP/PKA/CREB) pathway after binding to TSH receptors (TSHR) in the liver. The hepatic cAMP/PKA/CREB pathway also plays an important role in maintaining fasting glucose homeostasis. These findings implied a possible role for TSH in hepatic glucose metabolism. In this study, we used TSH receptor knockout mice (Tshr-ko mice) to clarify the effect of Tshr deletion on hepatic glucose metabolism, and investigated whether the effects of TSH directly regulate hepatic gluconeogenesis in HepG2 cells. Tshr-ko mice exhibited decreased fasting blood glucose levels, increased insulin sensitivity but normal level of fasting plasma insulin. Tshr deletion impaired hepatic glucose production by down-regulating the expression of glucose-6-phosphatase (G6P) and phosphoenolpyruvate pyruvate carboxylase (PEPCK) mRNA, two rate-limiting enzymes in hepatic gluconeogenesis, and enhancing the abundance of hepatic glucokinase (GK), the first enzyme regulating glycogen synthesis. Moreover, Tshr deletion inhibited the protein expression of hepatic phospho-CREB and increased the protein expression of hepatic phospho-AMP-activated protein kinase (p-AMPK), two up-stream regulators of PEPCK and G6P mRNA. In HepG2 cells, TSH increased the expression of G6P and PEPCK at mRNA level. These results indicated the simulative effects of TSH on hepatic glucose production in vivo and in vitro, suggesting a novel role for TSH in hepatic glucose metabolism.
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http://dx.doi.org/10.1507/endocrj.ej12-0462DOI Listing
March 2014

Akt and mTORC1 have different roles during liver tumorigenesis in mice.

Gastroenterology 2013 May 30;144(5):1055-65. Epub 2013 Jan 30.

Department of Surgery, University of Washington, Seattle, WA 98195, USA.

Background & Aims: Phosphatidylinositide 3-kinase (PI3K) is deregulated in many human tumor types, including primary liver malignancies. The kinase v-akt murine thymoma viral oncogene homolog 1 (Akt) and mammalian target of rapamycin complex (mTORC1) are effectors of PI3K that promote cell growth and survival, but their individual roles in tumorigenesis are not well defined.

Methods: In livers of albumin (Alb)-Cre mice, we selectively deleted tuberous sclerosis (Tsc)1, a negative regulator of Ras homolog enriched in brain and mTORC1, along with Phosphatase and tensin homolog (Pten), a negative regulator of PI3K. Tumor tissues were characterized by histologic and biochemical analyses.

Results: The Tsc1fl/fl;AlbCre, Ptenfl/fl;AlbCre, and Tsc1fl/fl;Ptenfl/fl;AlbCre mice developed liver tumors that differed in size, number, and histologic features. Livers of Tsc1fl/fl;AlbCre mice did not develop steatosis; tumors arose later than in the other strains of mice and were predominantly hepatocellular carcinomas. Livers of the Ptenfl/fl;AlbCre mice developed steatosis and most of the tumors that formed were intrahepatic cholangiocarcinomas. Livers of Tsc1fl/fl;Ptenfl/fl;AlbCre formed large numbers of tumors, of mixed histologies, with the earliest onset of any strain, indicating that loss of Tsc1 and Pten have synergistic effects on tumorigenesis. In these mice, the combination of rapamycin and MK2206 was more effective in reducing liver cell proliferation and inducing cell death than either reagent alone. Tumor differentiation correlated with Akt and mTORC1 activities; the ratio of Akt:mTORC1 activity was high throughout the course of intrahepatic cholangiocarcinomas development and low during hepatocellular carcinoma development. Compared with surrounding nontumor liver tissue, tumors from all 3 strains had increased activities of Akt, mTORC1, and mitogen-activated protein kinase and overexpressed fibroblast growth factor receptor 1. Inhibition of fibroblast growth factor receptor 1 in Tsc1-null mice suppressed Akt and mitogen-activated protein kinase activities in tumor cells.

Conclusions: Based on analyses of knockout mice, mTORC1 and Akt have different yet synergistic effects during the development of liver tumors in mice.
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http://dx.doi.org/10.1053/j.gastro.2013.01.053DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3633657PMC
May 2013