Associations of global biomarkers of oxidative stress with osteoporosis, bone microstructure and bone turnover: Evidence from human and animal studies.

PURPOSE: Human evidence on the association between oxidative stress and osteoporosis is inconsistent. Fluorescent Oxidation Products (FlOPs) are global biomarkers of oxidative stress. We examined the associations of FlOPs (excitation/emission wavelengths 320/420 nm for FlOP_320, 360/420 nm for FlOP_360, and 400/475 nm for FlOP_400) with osteoporosis, bone microstructure, and bone turnover markers in humans and rats. METHODS: In humans, we conducted a 1:2 age, sex, hospital, and specimen-matched case-control study to test the association between FlOPs and osteoporosis diagnosed from dual-energy X-ray absorptiometry. In eight-week-old male Wistar rats, we administrated D-galactose and 0.9 % saline for 90 days in treatment and control groups (n = 8/group); micro-CT was used to determine bone microstructure. RESULTS: In humans, higher levels of FlOP_320 (OR for per 1 SD increase = 1.49, 95 % CI: 1.01-2.20) and FlOP_360 (OR for per 1 SD increase = 1.59, 95 % CI: 1.07-2.37) were associated with increased odds of osteoporosis. FlOP_400 were not associated with osteoporosis. D-galactose treated rats, as compared with control rats, showed higher levels of FlOP_320 and MDA, and lower P1NP levels during 90 days of experiment (all P < 0.05). The D-galactose group had lower trabecular bone volume fraction (0.07 ± 0.03 vs. 0.13 ± 0.05; P = 0.008) and volumetric BMD (225.4 ± 13.8 vs. 279.1 ± 33.2 mg HA/cm3; P = 0.001) than the control group. CONCLUSION: In conclusion, higher FlOP_320 levels were associated with increased odds of osteoporosis, impaired bone microstructure and decreased bone formation.

Farnesyl diphosphate synthase promotes cell proliferation by regulating gene expression and alternative splicing profiles in HeLa cells.

Farnesyl diphosphate synthase (FDPS), an essential enzyme involved in the mevalonate pathway, is implicated in various diseases, including multiple types of cancer. As an RNA-binding protein (RBP), FDPS is also involved in transcriptional and post-transcriptional regulation. However, to the best of our knowledge, transcriptome-wide targets of FDPS still remain unknown. In the present study, FDPS expression patterns in pan-cancer were analyzed. In addition, it was investigated how FDPS overexpression (FDPS-OE) regulates the transcriptome in HeLa cells. FDPS-OE increased the proliferation rate in HeLa cells by MTT assay. Using transcriptome-wide high throughput sequencing and bioinformatics analysis, it was found that FDPS upregulated the expression levels of genes enriched in cell proliferation and extracellular matrix organization, including the laminin subunit γ2, interferon-induced proteins with tetratricopeptide repeats 2 and matrix metallopeptidase 19 genes. According to alternative splicing (AS) analysis, FDPS modulated the splicing patterns of the bone morphogenic protein 1, semaphorin 4D, annexin A2 and sirtuin 2 genes, which are enriched in the cell cycle and DNA repair, and are related to cell proliferation. To corroborate the FDPS-regulated transcriptome findings, FDPS was overexpressed in human osteosarcoma cells. Differentially expressed genes and regulated AS genes in the cells were both validated by reverse transcription-quantitative PCR. The results suggested that, as an emerging RBP, FDPS may serve an important role in transcriptome profiles by altering gene expression and regulating AS. FDPS also affected the cell proliferation rate. These findings broaden the understanding of the molecular functions of FDPS, and the potential of FDPS as a target in therapy should be investigated.

Atomically Interfacial Engineering on Molybdenum Nitride Quantum Dots Decorated N-doped Graphene for High-Rate and Stable Alkaline Hydrogen Production.

The development of low-cost, high-efficiency, and stable electrocatalysts for hydrogen evolution reaction (HER) under alkaline conditions is a key challenge in water electrolysis. Here, an interfacial engineering strategy that is capable of simultaneously regulating nanoscale structure, electronic structure, and interfacial structure of Mo2 N quantum dots decorated on conductive N-doped graphene via codoping single-atom Al and O (denoted as AlO@Mo2 N-NrGO) is reported. The conversion of Anderson polyoxometalates anion cluster ([AlMo6 O24 H6 ]3- , denoted as AlMo6) to Mo2 N quantum dots not only result in the generation of more exposed active sites but also in situ codoping atomically dispersed Al and O, that can fine-tune the electronic structure of Mo2 N. It is also identified that the surface reconstruction of AlOH hydrates in AlO@Mo2 N quantum dots plays an essential role in enhancing hydrophilicity and lowering the energy barriers for water dissociation and hydrogen desorption, resulting in a remarkable alkaline HER performance, even better than the commercial 20% Pt/C. Moreover, the strong interfacial interaction (MoN bonds) between AlO@Mo2 N and N-doped graphene can significantly improve electron transfer efficiency and interfacial stability. As a result, outstanding stability over 300 h at a current density higher than 100 mA cm-2 is achieved, demonstrating great potential for the practical application of this catalyst.

Dorzagliatin in drug-naïve patients with type 2 diabetes: a randomized, double-blind, placebo-controlled phase 3 trial.

Improving glucose sensitivity remains an unmet medical need in treating type 2 diabetes (T2D). Dorzagliatin is a dual-acting, orally bioavailable glucokinase activator that enhances glucokinase activity in a glucose-dependent manner, improves glucose-stimulated insulin secretion and demonstrates effects on glycemic control in patients with T2D. We report the findings of a randomized, double-blind, placebo-controlled phase 3 clinical trial to evaluate the efficacy and safety of dorzagliatin in patients with T2D. Eligible drug-naïve patients with T2D (n = 463) were randomly assigned to the dorzagliatin or placebo group at a ratio of 2:1 for 24 weeks of double-blind treatment, followed by 28 weeks of open-label treatment with dorzagliatin for all patients. The primary efficacy endpoint was the change in glycated hemoglobin from baseline to week 24. Safety was assessed throughout the trial. At week 24, the least-squares mean change in glycated hemoglobin from baseline (95% confidence interval) was -1.07% (-1.19%, -0.95%) in the dorzagliatin group and -0.50% (-0.68%, -0.32%) in the placebo group (estimated treatment difference, -0.57%; 95% confidence interval: -0.79%, -0.36%; P < 0.001). The incidence of adverse events was similar between the two groups. There were no severe hypoglycemia events or drug-related serious adverse events in the dorzagliatin group. In summary, dorzagliatin improved glycemic control in drug-naïve patients with T2D and showed a good tolerability and safety profile.

The Differential Expression of Circular RNAs in Type 2 Diabetes Mellitus and Latent Autoimmune Diabetes in Adults.

Objective: Expression of circular RNAs (circRNAs) in the peripheral blood of individuals with latent autoimmune diabetes in adults (LADA) and type 2 diabetes mellitus (T2DM) were quantified to identify dysregulated circRNAs compared with control individuals. Methods: circRNAs were obtained from the peripheral blood serum of 12 healthy adults and 12 individuals with LADA and 12 type 2 diabetics. The circRNA expression profiles were analyzed by high-throughput RNA sequencing. The most highly dysregulated circular RNAs were validated by quantitative real-time polymerase chain reaction. A circular RNA-microRNA (miRNA) network diagram predicted the interactions of circular RNAs, miRNAs, and coding genes. Results: A total of 2334 differentially expressed circRNAs were detected among the three groups, with 277 circRNAs in the Group DM versus Group NG; 992 circRNAs in the Group LADA versus Group NG and 1065 circRNAs in the Group DM versus Group LADA. Six circRNAs were identified as the most distinctive differentially expressed targets (p < 0.05). The proposed molecular functions of these differentially expressed circRNAS included the tumor necrosis factor signaling pathway, the FoxO signaling pathway, cellular senescence, and long-term potentiation (all false discovery rate p < 0.05) which may contribute to T2DM and LADA. Conclusion: circRNAs are aberrantly expressed in the peripheral blood of patients with T2DM and LADA and may interact with miRNA and circRNA-derived peptides in the development of diabetes. Further investigations may illustrate the partial pathogenesis of diabetes mellitus. Clinical Trial Registration number: ChiCTR1900020644.

Full recovery from chronic headache and hypopituitarism caused by lymphocytic hypophysitis: A case report.

BACKGROUND: Lymphocytic hypophysitis (LYH) is an important condition to consider in the differential diagnosis of patients with a pituitary mass. The main clinical manifestations of LYH include headache, symptoms related to sellar compression, hypopituitarism, diabetes insipidus and hyperprolactinemia. Headache, which is a frequent complaint of patients with LYH, is thought to be related to the occupying effect of the pituitary mass and is rapidly resolved with a good outcome after timely and adequate glucocorticoid treatment or surgery. CASE SUMMARY: Here, we report a patient with LYH whose initial symptom was headache and whose pituitary function assessment showed the presence of secondary hypoadrenalism, central hypothyroidism and hypogonadotropic hypogonadism. Pituitary magnetic resonance imaging showed symmetrical enlargement of the pituitary gland with suprasellar extension in a dumbbell shape with significant homogeneous enhancement after gadolinium enhancement. The size of the gland was approximately 17.7 mm × 14.3 mm × 13.8 mm. The pituitary stalk was thickened without deviation, and there was an elevation of the optimal crossing. The lesion grew bilaterally toward the cavernous sinuses, and the parasternal dural caudal sign was visible. The patient presented with repeatedly worsening and prolonged headaches three times even though the hypopituitarism had fully resolved after glucocorticoid treatment during this course. CONCLUSION: This rare headache regression suggests that patients with chronic headaches should also be alerted to the possibility of LYH.

Plasma Fluorescent Oxidation Products and Bone Mineral Density Among Male Veterans: A Cross-Sectional Study.

In vitro and vivo studies indicate that oxidative stress contributes to bone loss. Fluorescent oxidation products (FlOPs) are novel biomarkers of oxidative stress; they reflect global oxidative damage of lipids, proteins, carbohydrates, and DNA. However, whether FlOPs are associated with bone mineral density (BMD) is still unclear. In the present study, we examined the association between FlOPs and BMD among male veterans. This cross-sectional study was conducted among participants recruited from the Department of Medical Examination, The Second Hospital of Jilin University in Jilin, China. We identified male veterans who were at least 50 y old between June and October of 2019. Plasma FlOPs were measured with a fluorescent microplate reader (excitation/emission wavelength: 320/420 nm). BMD were measured by dual-energy X-ray absorptiometry (DXA). The association between FlOPs and BMD was tested by multivariable linear regression models. A total of 164 male veterans were enrolled in the study, the average age was 56.6 y. After adjusting for covariates, veterans who had FlOP levels in the highest tertile had a statistically significant lower femoral neck (ß = -0.044; p = 0.007) and total hip BMD (ß = -0.045; p = 0.020) as compared to those with FlOP levels in the lowest tertile. Similar results were found when FlOPs were treated as a continuous variable (per 1-SD increase, ß = -0.014 and p = 0.033 for femoral neck BMD; ß = -0.016 and p = 0.047 for total hip BMD). Higher FlOP levels were associated with lower BMD among male veterans.

Association between lipid biomarkers and osteoporosis: a cross-sectional study.

BACKGROUND: Osteoporosis and cardiovascular diseases (CVDs) are 2 major public health problems. Osteoporosis and CVDs may be linked but the association between lipid profile and osteoporosis is still controversial. The purpose of this study was to examine the associations of total cholesterol (TC), low-density lipoprotein cholesterol (LDL-C), high-density lipoprotein cholesterol (HDL-C) and triglyceride (TG) with osteoporosis. METHODS: Using inpatients' and outpatients' electronic medical records (EMR) and dual X-ray absorptiometry (DXA) database stored at The Second Hospital of Jilin University, we included 481 individuals with complete and valid lipid and bone mineral density (BMD) data in 2017. Serum samples were used to measure TC, LDL-C, HDL-C and TG. Femoral neck and total hip BMD were measured by DXA; osteoporosis was defined as femoral neck or total hip T-score ≤ -2.5. Multivariable logistic regression models were used to test the associations of TC, LDL-C, HDL-C and TG with osteoporosis. RESULTS: The mean age for included individuals was 62.7 years (SD = 8.6 years); 60.1 % of them were female. Each standard deviation (SD) increase in TC (Odds Ratio [OR]: 1.48; 95 % Confidence Interval [CI]: 1.06-2.07) and TG (OR: 1.67; 95 % CI: 1.16-2.39) were associated with increased risk of osteoporosis; LDL-C and HDL-C levels were not associated with osteoporosis. Age, sex and body mass index (BMI) did not interact with the relationships of TC and TG with osteoporosis (all P > 0.10). CONCLUSIONS: Higher TC and TG levels were associated with greater risk of osteoporosis in this cross-sectional study.

Severe metabolic disorders coexisting with Werner syndrome: a case report.

Werner syndrome, also called adult progeria, is a heritable autosomal recessive human disorder characterized by the premature onset of numerous age-related diseases including juvenile cataracts, dyslipidemia, diabetes mellitus (DM), osteoporosis, atherosclerosis, and cancer. Werner syndrome is a segmental progeroid syndrome whose presentation resembles accelerated aging. The most common causes of death for WS patients are atherosclerosis and cancer. A 40-year-old female presented with short stature, bird-like facies, canities with alopecia, scleroderma-like skin changes, and non-healing foot ulcers. The patient reported a history of delayed puberty, abortion, hypertriglyceridemia, and juvenile cataracts. A clinical diagnosis of WS was made and subsequently confirmed. We discovered two WRN gene mutations in the patient, Variant 1 was the most common WRN mutation, nonsense mutation (c.1105C>T:p.R369Ter) in exon 9, which caused a premature termination codon (PTC) at position 369. Variant 2 was a frameshift mutation (c.1134delA:p.E379KfsTer5) in exon 9, which caused a PTC at position 383 and has no published reports describing. Patients with WS can show a wide variety of clinical and biological manifestations in endocrine-metabolic systems (DM, thyroid dysfunction, and hyperlipidemia). Doctors must be cognizant of early manifestations of WS and treatment options.

Efficacy and safety of chiglitazar, a novel peroxisome proliferator-activated receptor pan-agonist, in patients with type 2 diabetes: a randomized, double-blind, placebo-controlled, phase 3 trial (CMAP).

Chiglitazar (Carfloglitazar) is a novel non-thiazolidinedione (TZD) structured peroxisome proliferator-activated receptor (PPAR) pan-agonist that has shown promising effects on glycemic control and lipid regulation in patients with type 2 diabetes in previous clinical studies. This randomized phase 3 trial aimed to compare the efficacy and safety of chiglitazar with placebo in patients with type 2 diabetes with insufficient glycemic control by strict diet and exercise alone. Eligible patients were randomly assigned to receive chiglitazar 32 mg (n = 167), chiglitazar 48 mg (n = 166), or placebo (n = 202) once daily. The primary endpoint was the change in glycosylated hemoglobin A1c (HbA1c) at week 24 with superiority of chiglitazar over placebo. The results showed that both chiglitazar 32 and 48 mg resulted in significant and clinically meaningful reductions in HbA1c, and placebo-adjusted estimated treatment differences at week 24 for chiglitazar 32 and 48 mg were -0.87% (95% confidential interval (CI): -1.10 to -0.65; P < 0.0001) and -1.05% (95% CI: -1.29 to -0.81; P < 0.0001), respectively. Secondary efficacy parameters including glycemic control, insulin sensitivity and triglyceride reduction were also significantly improved in the chiglitazar groups. The overall frequency of adverse events and study discontinuation attributable to adverse events were similar among the groups. Low incidences of mild edema and body weight gain were reported in the chiglitazar dose groups. The results from this phase 3 trial demonstrated that the PPAR pan-agonist chiglitazar possesses an overall good efficacy and safety profile in patients with type 2 diabetes inadequately controlled with lifestyle interventions, thereby providing adequate supporting evidence for using this PPAR pan-agonist as a treatment option for type 2 diabetes.

Differential Profile of Plasma Circular RNAs in Type 1 Diabetes Mellitus.

BACKGROUND: No currently available biomarkers or treatment regimens fully meet therapeutic needs of type 1 diabetes mellitus (T1DM). Circular RNA (circRNA) is a recently identified class of stable noncoding RNA that have been documented as potential biomarkers for various diseases. Our objective was to identify and analyze plasma circRNAs altered in T1DM. METHODS: We used microarray to screen differentially expressed plasma circRNAs in patients with new onset T1DM (n=3) and age-/gender-matched healthy controls (n=3). Then, we selected six candidates with highest fold-change and validated them by quantitative real-time polymerase chain reaction in independent human cohort samples (n=12). Bioinformatic tools were adopted to predict putative microRNAs (miRNAs) sponged by these validated circRNAs and their downstream messenger RNAs (mRNAs). Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway analyses were performed to gain further insights into T1DM pathogenesis. RESULTS: We identified 68 differentially expressed circRNAs, with 61 and seven being up- and downregulated respectively. Four of the six selected candidates were successfully validated. Curations of their predicted interacting miRNAs revealed critical roles in inflammation and pathogenesis of autoimmune disorders. Functional relations were visualized by a circRNA-miRNA-mRNA network. GO and KEGG analyses identified multiple inflammation-related processes that could be potentially associated with T1DM pathogenesis, including cytokine-cytokine receptor interaction, inflammatory mediator regulation of transient receptor potential channels and leukocyte activation involved in immune response. CONCLUSION: Our study report, for the first time, a profile of differentially expressed plasma circRNAs in new onset T1DM. Further in silico annotations and bioinformatics analyses supported future application of circRNAs as novel biomarkers of T1DM.

Let-7b downgrades CCND1 to repress osteogenic proliferation and differentiation of MC3T3-E1 cells: An implication in osteoporosis.

The aim of this study was to reveal the effect of let-7b on osteoporosis (OP). Synthetic let-7b mimics or inhibitors were transfected into MC3T3-E1 cells. The expression of let-7b in MC3T3-E1 and its effect on cell viability, apoptosis, and the apoptosis-related proteins (Bcl-2, Bax, and cleaved caspase-9) were tested by CCK-8 assay, flow cytometry and Western blot, severally. The osteogenic differentiation markers (Runx2 and Osterix) and Wnt/ß-catenin pathway related markers (ß-catenin and C-myc) were detected by qRT-PCR and Western blot. The relationships between let-7b and cyclin D1 (CCND1) were confirmed by luciferase reporter assay. The differentiation and mineralization of MC3T3-E1 cells were analyzed by alkaline phosphatase (ALP) activity assay and alizarin red staining. The outcomes indicated that overexpression/ablation of let-7b repressed/facilitated MC3T3-E1 cell viability and accelerated/suppressed MC3T3-E1 cell apoptosis. Besides, a remarkable decrease/augment of Bcl-2 protein expression and the distinct fortify/reduction of Bax and cleaved caspase-9 expression levels were observed in let-7b mimics/inhibitors group in MC3T3-E1 cells. Moreover, we discovered that let-7b overexpression/ablation retrained/facilitated the mRNA and protein expression of Runx2 and Osterix. It was confirmed that CCND1 was a downstream target of let-7b and was negatively modulated by let-7b. In addition, high-expression/deficiency of let-7b inhibited/increased the expression levels of ß-catenin and C-myc in MC3T3-E1 cells. Taken together, our study revealed that let-7b overexpression/depletion repressed/accelerated MC3T3-E1 cell proliferation, differentiation, and mineralization while promoted/suppressed MC3T3-E1 cell apoptosis through targeting CCND1, which might be adjusted by Wnt/ß-catenin pathway. Our findings might offer a basis for developing novel targets for OP treatment.

miR-1258: a novel microRNA that controls TMPRSS4 expression is associated with malignant progression of papillary thyroid carcinoma.

BACKGROUND: MicroRNA-1258 (miR-1258) has been shown to play an anti-cancer role in a variety of cancers, but its relationship with papillary thyroid cancer (PTC) has not been reported. The emphasis of this research was to reveal the biological function of miR-1258 in PTC and its potential mechanisms. MATERIAL AND METHODS: We measured miR-1258 expression in PTC cells and the transfection efficiency of miR-1258 mimic and miR-1258 inhibitor by quantitative real-time PCR (qRT-PCR) assay. Cell Counting Kit-8 assay (CCK8) and Transwell experiments were conducted to examine the influences of altering miR-1258 expression on the viability, migration, and invasion of PTC cells. Bioinformatics prediction and dual-luciferase experiment were performed to verify the target gene of miR-1258. Finally, we carried out a rescue assay to verify whether the regulation of miR-1258 on the biological behaviour of PTC cells needs to be achieved by regulating TMPRSS4. RESULTS: The outcomes revealed that miR-1258 was lowly expressed in PTC cell lines and miR-1258 showed the lowest expression in KTC-1 and the highest expression in B-CPAP among all tested PTC cell lines. Overexpression of miR-1258 inhibited KTC-1 cell viability and ability to migrate and invade, whereas inhibition of miR-1258 in B-CPAP cells has the opposite effect. Furthermore, we affirmed that miR-1258 can directly target TMPRSS4, and miR-1258 can reduce the biological malignant behaviour of PTC cells via regulation of TMPRSS4. CONCLUSION: Taken together, our research raised the possibility that miR-1258 was an anti-oncogene, which exerts its anti-cancer function by targeting TMPRSS4. Hence, it may be possible to treat PTC by targeting the miR-1258/TMPRSS4 axis in the future.

Interleukin-10 Protects Schwann Cells against Advanced Glycation End Products-Induced Apoptosis via NF-κB Suppression.

Demyelination resulting from Schwann cell injury is a main pathological feature of diabetic neuropathy, and a key contributor to this process may be inflammation due to advanced glycation end products (AGEs). Therefore, protection by anti-inflammation agents is anticipated. In this study, we showed that interleukin-10 (IL-10), an anti-inflammatory cytokine, inhibits apoptosis of Schwann cells induced by AGEs in vitro. We isolated and cultured Schwann cells from rat sciatic nerves. As detected by flow cytometry, apoptosis of Schwann cells markedly increased following incubation with AGEs for 48 h. However, pretreatment with IL-10 inhibited AGE-induced apoptosis. The effect of IL-10 on NF-κB, which is a very important regulator of inflammation, was also evaluated, and results showed high levels of phospho-NF-κB and nuclear localization of NF-κB in cells incubated with AGEs but low levels of phospho-NF-κB and cytoplasmic localization in the cells incubated with IL-10, indicating the activation of NF-κB by AGEs and inhibition of NF-κB by IL-10. Moreover, incubating Schwann cells with an NF-κB inhibitor (caffeic acid phenethyl ester) for 30 min before adding AGEs mimicked IL-10, lowering the amount of reactive oxygen species and activity of caspase-3 and also decreasing apoptosis in Schwann cells. These results indicate that IL-10 may protect Schwann cells against AGE-induced apoptosis by attenuating oxidative stress via the inhibition of activation of NF-κB.

miR-218 regulates diabetic nephropathy via targeting IKK-ß and modulating NK-κB-mediated inflammation.

Diabetic nephropathy (DN) is a common clinically relevant complication of diabetes that is associated with damage to the capillaries, yet the etiology of this condition remains unclear. Nuclear factor-kappa B (NF-κB) activation is known to be associated with DN-related inflammation and disease progression. Recent work indicated that microRNAs are diagnostic biomarkers of DN progression associated with inflammation in the progression of DN. miR-218 is known to play key regulatory roles in certain cancers in humans, while its influence on DN pathology remains uncertain. The present study, therefore, sought to assess how miR-218 influences the progression of disease in both a rat streptozotocin-induced model of DN and as well as an in vitro model system in which mouse podocytes were stimulated with high glucose levels. We found miR-218 to be markedly downregulated in both model systems relative to appropriate controls, and this downregulation was associated with IKK-ß upregulation. In DN rat model, overexpressing miR-218 was sufficient to reduce renal injury. We further determined that podocyte proliferation was markedly impaired by glucose treatment, leading to the apoptotic death of these cells, and miR-218 mimics were able to reduce these phenotypes. Overexpressing miR-218 also significantly dampened inflammatory responses in this model system, as evidenced by reduced tumor necrosis factor-α, interleukin-6 (IL-6), IL-1ß, and MCP-1 levels. We then confirmed that miR-218 targeting the messenger RNA encoding IKK-ß using a dual-luciferase reporter assay. Together, our results provide clear evidence that miR-218 regulate NF-κB-mediated inflammation, which is central to DN progression.

1,25-(OH)2D3 protects Schwann cells against advanced glycation end products-induced apoptosis through PKA-NF-κB pathway.

AIMS: To explore the effect and mechanism of 1, 25-(OH)2D3 on Schwann cell apoptosis induced by advanced glycation end products. MAIN METHODS: Schwann cells, isolated from rodent sciatic nerve were incubated with AGE-modified bovine serum albumin(AGE) to mimic diabetic conditions and 1,25-(OH)2D3 was used as protector. Cell apoptosis was detected by PI/Annexin-V staining, caspase 3 activity assay and western blotting for caspase 3 and PARP. The activation of protein kinase A (PKA) and nuclear factor kappa-B (NF-κB) was evaluated by western blot. Immunofluorescent staining was used for intercellular location of NF-κB. Cytokine secretion was evaluated by enzyme-linked immunosorbent assay. KEY FINDINGS: Schwann cell apoptosis accelerated after incubating with AGE. However, if combining 1,25-(OH)2D3 with AGE, apoptosis decreased significantly. 1,25-(OH)2D3 enhanced PKA activity, but inhibited AGE-induced nuclear translocation of NF-κB. Furthermore, PKA activator (8-bromoadenoside cyclic adenoside monophosphate, 8-Br-cAMP) or NF-κB inhibitor (caffeic acid phenethyl ester, CAPE) could reduce the apoptosis, decreased cleaved caspase 3 and cleaved PARP, suggesting the involvement of PKA and NF-κB pathways in the protection of 1,25-(OH)2D3 on Schwann cells. Moreover, 8-Br-cAMP and CAPE could inhibit AGE-induced secretion of interleukin(IL)-1ß, prostaglandin E2(PEG2) and cyclooxygenase 2(COX2). Interestingly, 8-Br-cAMP decreased phospho-NF-κB and inhibited nucleus translocation of NF-κB. It hinted at the regulation of PKA to NF-κB. Finally, a pre-treatment of H-89 (an inhibitor of PKA) could block the protection of 1,25-(OH)2D3 on cell apoptosis. In conclusion, 1,25-(OH)2D3 could protect Schwann cell against AGE-induced apoptosis through PKA/NF-κB pathway. SIGNIFICANCE: These findings provide experimental rationales for using vitamin D for diabetic neuropathy.

Celiac Disease Autoimmunity in Patients with Autoimmune Diabetes and Thyroid Disease among Chinese Population.

The prevalence of celiac disease autoimmunity or tissue transglutaminase autoantibodies (TGA) amongst patients with type 1 diabetes (T1D) and autoimmune thyroid disease (AITD) in the Chinese population remains unknown. This study examined the rate of celiac disease autoimmunity amongst patients with T1D and AITD in the Chinese population. The study included 178 patients with type 1 diabetes and 119 with AITD where 36 had both T1D and AITD, classified as autoimmune polyglandular syndrome type 3 variant (APS3v). The study also included 145 patients with type 2 diabetes (T2D), 97 patients with non-autoimmune thyroid disease (NAITD), and 102 healthy controls. Serum islet autoantibodies, thyroid autoantibodies and TGA were measured by radioimmunoassay. TGA positivity was found in 22% of patients with either type 1 diabetes or AITD, much higher than that in patients with T2D (3.4%; p< 0.0001) or NAITD (3.1%; P < 0.0001) or healthy controls (1%; p<0.0001). The patients with APS3v having both T1D and AITD were 36% positive for TGA, significantly higher than patients with T1D alone (p = 0.040) or with AITD alone (p = 0.017). T1D and AITD were found to have a 20% and 30% frequency of overlap respectively at diagnosis. In conclusion, TGA positivity was high in the Chinese population having existing T1D and/or AITD, and even higher when both diseases were present. Routine TGA screening in patients with T1D or AITD will be important to early identify celiac disease autoimmunity for better clinical care of patients.

Evaluation of islets derived from human fetal pancreatic progenitor cells in diabetes treatment.

INTRODUCTION: With the shortage of donor organs for islet transplantation, insulin-producing cells have been generated from different types of stem cell. Human fetal pancreatic stem cells have a better self-renewal capacity than adult stem cells and can readily differentiate into pancreatic endocrine cells, making them a potential source for islets in diabetes treatment. In the present study, the functions of pancreatic islets derived from human fetal pancreatic progenitor cells were evaluated in vitro and in vivo. METHODS: Human pancreatic progenitor cells isolated from the fetal pancreas were expanded and differentiated into islet endocrine cells in culture. Markers for endocrine and exocrine functions as well as those for alpha and beta cells were analyzed by immunofluorescent staining and enzyme-linked immunosorbent assay (ELISA). To evaluate the functions of these islets in vivo, the islet-like structures were transplanted into renal capsules of diabetic nude mice. Immunohistochemical staining for human C-peptide and human mitochondrion antigen was applied to confirm the human origin and the survival of grafted islets. RESULTS: Human fetal pancreatic progenitor cells were able to expand in medium containing basic fibroblast growth factor (bFGF) and leukemia inhibitor factor (LIF), and to differentiate into pancreatic endocrine cells with high efficiency upon the actions of glucagon-like peptide-1 and activin-A. The differentiated cells expressed insulin, glucagon, glucose transporter-1 (GLUT1), GLUT2 and voltage-dependent calcium channel (VDCC), and were able to aggregate into islet-like structures containing alpha and beta cells upon suspension. These structures expressed and released a higher level of insulin than adhesion cultured cells, and helped to maintain normoglycemia in diabetic nude mice after transplantation. CONCLUSIONS: Human fetal pancreatic progenitor cells have good capacity for generating insulin producing cells and provide a promising potential source for diabetes treatment.

α-Cell loss from islet impairs its insulin secretion in vitro and in vivo.

Pancreatic islet transplantation is an effective treatment for diabetes mellitus. But it is not clear whether α-cell loss during islets isolation could impair the insulin release from ß-cell. To unravel this issue, human islets with α-cell deficiency were prepared by prolonged enzyme digestion, as confirmed by immunocytochemistry, immunofluorescence staining and islet insulin/glucagon content analysis. The functions of islets with α-cell deficiency were evaluated in vitro and in vivo. In vitro, human islets with α-cell deficiency exhibited low glucose-induced insulin release compared with intact islets. In islets deficient in α-cells, exogenous glucagon did not alone stimulate insulin release in the absence of glucose, but increased the glucose-induced insulin release in a dose-dependent manner. In intact islets, glucagon did not significantly change the glucose-stimulated insulin secretion. In vivo, transplantation of human islets with α-cell deficiency did not effectively correct hyperglycemia in diabetic C57BL/6 mice. In diabetic nude mice transplanted with islets deficient in α-cells, administration of exogenous glucagon significantly decreased glycemia, while withdrawing glucagon increased glycemic levels as compared with relevant controls. In addition, the survival of diabetic nude mice grafted with islets deficient in α-cells was significantly shorter than the survival of nude mice grafted with intact islets. These results indicated that glucagon-secreting α-cells have an important role in maintaining glucose-stimulated insulin release from ß-cells, and that α-cell loss from islets during isolation has a deleterious effect on insulin secretion.

Platelet adhesion and fusion to endothelial cell facilitate the metastasis of tumor cell in hypoxia-reoxygenation condition.

To investigate the relevant molecular mechanisms of platelet in promoting metastasis of tumor cell. The adhesion of fluorescence dye labeled-platelet to human liver sinusoidal endothelial cell (LSEC) line and tumor cell lines were detected by fluorescence microscope and fluorescence plate reader or laser scanning confocal microscope. The relevant adhesion molecules were analyzed by the antibody blockage experiment. The immune colloidal gold transmission electron microscope (TEM), flow cytometry and dye transfer were used to decipher the adhesion and fusion of platelet and LSEC. The tumor cells adhesion to vessels in ischemia condition was analyzed on mouse mesenteric vessels and the metastasis and neovascularization of metastatic foci in pulmonary tissue were also detected after tumor cells injected into nude mice via tail veil. After hypoxia-reoxygenation, tumor cell or LSEC markedly increased its adhesion with platelet, which could be blocked by different antibodies to platelet adhesion molecules. Platelet increased adhesion of tumor cell to LSEC in dose-dependent manner. The fusion of platelet and LSEC was demonstrated by translocation of fluorescent dye from platelet into the adherent LSEC; gpIIb emerged on the LSEC; and confirmed by TEM. The morphological examination found platelet presented between tumor cell and LSEC. Animal experiment indicated that the tumor adhesion to vessels was seldom in normal condition, but increased in ischemia-reperfusion condition, and further significantly enhanced by platelets. The number of tumor metastatic foci and the density of blood vessels within metastatic foci in lung were markedly increased by tumor cell pre-adhered with platelet. The adhesion or fusion of platelet to endothelial cell mediated by platelet surface adhesion molecules, which could promote the adhesion of tumor cell with endothelial cells and the tumor metastasis.