Association between HIF-1α, BNIP3, and autophagy in the chorionic villi of missed abortion.

AIM: To investigate the expression of autophagy mediated by the hypoxia-inducible factor 1α (HIF-1α)/BNIP3 signaling pathway in villus tissues of missed abortion and HTR-8/SVneo cells and to elucidate the association of HIF-1α and BNIP3 in autophagy of missed abortion. METHODS: Villus tissues from 30 healthy women with induced abortion and 35 patients with missed abortion were collected, and HTR-8/SVneo cells were cultured under hypoxia and transfected with HIF-1α-siRNA. Real-time polymerase chain reaction was utilized to measure the mRNA levels of HIF-1α and BNIP3; Western blotting was performed to determine the protein levels of HIF-1α, BNIP3, LC3 II/I, and Beclin 1 in villus tissues and HTR-8/SVneo cells. Cellular invasion activity was detected by transwell matrigel assay. The level of autophagy was confirmed by transmission electron microscopy of autophagosome formation. RESULTS: The mRNA levels of HIF-1α and BNIP3 were significantly lower in the missed abortion villi than in the induced abortion samples. The protein levels of HIF-1α, BNIP3, Beclin 1, and LC3II/I were significantly decreased in villus tissues from missed abortion, and autophagosomes were significantly decreased in villus tissues from missed abortion. Under hypoxia, the mRNA expression of HIF-1α and BNIP3 was inhibited after silencing HIF-1α by RNAi, while the protein expression of HIF-1α, BNIP3, Beclin1, and LC3II/I was significantly downregulated. The number of invading cells was significantly decreased, and autophagosomes were significantly decreased after silencing HIF-1α by RNAi in HTR-8/SVneo cells. CONCLUSIONS: Autophagy mediated by the HIF-1α/BNIP3 signaling pathway in villous trophoblast cells may be associated with the progression and development of missed abortion.

TRAF4, a new substrate of SIAH1, participates in chemotherapy resistance of breast cancer cell by counteracting SIAH1-mediated downregulation of ß-catenin.

PURPOSE: TRAF4 plays an important role in the development and progression of breast cancer, but its impact on chemotherapy resistance is as yet, however, poorly understood. METHODS: Western blotting, immunoprecipitation, and immunofluorescence staining were used to identify and verify that TRAF4 was a novel substrate of SIAH1 and prevented SIAH1-mediated ß-catenin degradation. Cell proliferation analysis and Flow cytometry analysis were utilized to detect TRAF4's function on the growth-inhibitory effect of etoposide. Immunohistochemistry was used to detect the expression of TRAF4, SIAH1, and ß-catenin. Statistical analysis was used to analyze the relationships between them with clinical parameters and curative effect of chemotherapy pathologically. RESULTS: Our results suggested that TRAF4 prevents SIAH1-mediated ß-catenin degradation. TRAF4 was a novel substrate of SIAH1 and the TRAF domain of TRAF4 was critical for binding to SIAH1. TRAF4 reduced the growth-inhibitory effect of etoposide via reducing the number of S-phase cells and suppressing cell apoptosis. Concordantly, we found that breast cancer patients with a low-TRAF4 expression benefited most from chemotherapy, who had higher tumor volume reduction rate and better pathological response, while, the high-TRAF4 expression group had lower tumor volume reduction rate and poor pathological response. CONCLUSIONS: TRAF4 was a novel substrate of SIAH1 and prevented SIAH1-mediated ß-catenin degradation, which explains the protective effect of TRAF4 on ß-catenin during cell stress and links TRAF4 to chemotherapy resistance in tumors. These findings implicated a novel pathway for the oncogenic function of TRAF4.

Feasibility of human hair follicle-derived mesenchymal stem cells/CultiSpher(®)-G constructs in regenerative medicine.

The use of human mesenchymal stem cells (hMSCs) in cell therapies has increased the demand for strategies that allow efficient cell scale-up. Preliminary data on the three-dimensional (3D) spinner culture describing the potential use of microcarriers for hMSCs culture scale-up have been reported. We exploited a rich source of autologous stem cells (human hair follicle) and demonstrated the robust in vitro long-term expansion of human hair follicle-derived mesenchymal stem cells (hHF-MSCs) by using CultiSpher(®)-G microcarriers. We analyzed the feasibility of 3D culture by using hHF-MSCs/CultiSpher(®)-G microcarrier constructs for its potential applicability in regenerative medicine by comparatively analyzing the performance of hHF-MSCs adhered to the CultiSpher(®)-G microspheres in 3D spinner culture and those grown on the gelatin-coated plastic dishes (2D culture), using various assays. We showed that the hHF-MSCs seeded at various densities quickly adhered to and proliferated well on the microspheres, thus generating at least hundreds of millions of hHF-MSCs on 1 g of CultiSpher(®)-G within 12 days. This resulted in a cumulative cell expansion of greater than 26-fold. Notably, the maximum and average proliferation rates in 3D culture were significantly greater than that of the 2D culture. However, the hHF-MSCs from both the cultures retained surface marker and nestin expression, proliferation capacity and differentiation potentials toward adipocytes, osteoblasts and smooth muscle cells and showed no significant differences as evidenced by Edu incorporation, cell cycle, colony formation, apoptosis, biochemical quantification and qPCR assays.

Effects of Adjuvant Radiation Plus Chemotherapy on Survival Outcomes in Stage III C Endometrial Cancer According to Histology: Analysis of Data from the Surveillance, Epidemiology, and End Results Database.

Purpose: To evaluate the survival benefit of radiation plus chemotherapy in adult females with stage IIIC endometrial cancer and to investigate whether the benefit varies according to histology. Methods: Data from adult females with International Federation of Gynecology and Obstetrics (FIGO) stage IIIC endometrial cancer, who underwent at least total hysterectomy between 2010 and 2015, were obtained from the Surveillance, Epidemiology, and End Results (SEER) database. Adjuvant treatments were categorized as chemotherapy alone, chemotherapy with external beam radiation therapy (EBRT), chemotherapy with vaginal brachytherapy (VBT), or chemotherapy with EBRT+VBT. Multivariate Cox regression models, Kaplan-Meier curves, and log-rank tests were used to assess the association between treatment modality and overall survival (OS). Results: In total, 2138 cases were identified: stage IIIC1 (n = 1299 [60.8%]) and stage IIIC2 (n = 839 [39.2%]). Median OS for all patients was 48 (interquartile range [IQR] 28-70) months. Regarding adjuvant treatment, 40.5% of patients underwent chemotherapy only, followed by chemotherapy with EBRT (35.5%). Stage IIIC patients treated with chemotherapy plus radiation exhibited a significantly reduced risk for death from endometrial cancer in both univariate and multivariate analyses (P < 0.001). However, when stratified according to histology, OS also differed according to treatment modality when analyzing each histological type; combination therapy was no longer significantly different from chemotherapy alone for any histology (clear cell and carcinosarcoma). Combination therapy was associated with improved OS in patients with IIIC1 and IIIC2 disease. Similar associations were observed in patients with high-grade stage IIIC endometrioids. However, for low-grade tumors, combination therapy was no longer associated with reduced risk for death compared with chemotherapy alone. Conclusion: For patients with stage IIIC endometrial cancer, combined treatment with radiation and chemotherapy was associated with improved OS compared with chemotherapy alone. However, no survival benefit was found, and radiotherapy may be unnecessary in patients with low-grade endometrioids.

Adalimumab ameliorates memory impairments and neuroinflammation in chronic cerebral hypoperfusion rats.

Vascular dementia (VaD) is the second most common type of dementia worldwide. Although there are five FDA-approved drugs for the treatment of Alzheimer's disease (AD), none of them have been applied to treat VaD. Adalimumab is a TNF-α inhibitor that is used for the treatment of autoimmune diseases such as rheumatoid arthritis. In a recent retrospective case-control study, the application of adalimumab for rheumatoid or psoriasis was shown to decrease the risk of AD. However, whether adalimumab can be used for the treatment of VaD is not clear. In this study, we used 2VO surgery to generate a VaD rat model and treated the rats with adalimumab or vehicle. We demonstrated that VaD rats treated with adalimumab exhibited significant improvements in memory. In addition, adalimumab treatment significantly alleviated neuronal loss in the hippocampi of VaD rats. Moreover, adalimumab significantly reduced microglial activation and reversed M1/M2 polarization in VaD rats. Furthermore, adalimumab treatment suppressed the activity of NF-κB, an important neuroinflammatory transcription factor. Finally, adalimumab displayed a protective role against oxidative stress in VaD rats. Our results indicate that adalimumab may be applied for the treatment of human patients with VaD.

Large-scale expansion of Wharton's jelly-derived mesenchymal stem cells on gelatin microbeads, with retention of self-renewal and multipotency characteristics and the capacity for enhancing skin wound healing.

INTRODUCTION: Successful stem cell therapy relies on large-scale generation of stem cells and their maintenance in a proliferative multipotent state. This study aimed to establish a three-dimension culture system for large-scale generation of hWJ-MSC and investigated the self-renewal activity, genomic stability and multi-lineage differentiation potential of such hWJ-MSC in enhancing skin wound healing. METHODS: hWJ-MSC were seeded on gelatin microbeads and cultured in spinning bottles (3D). Cell proliferation, karyotype analysis, surface marker expression, multipotent differentiation (adipogenic, chondrogenic, and osteogenic potentials), and expression of core transcription factors (OCT4, SOX2, NANOG, and C-MYC), as well as their efficacy in accelerating skin wound healing, were investigated and compared with those of hWJ-MSC derived from plate cultres (2D), using in vivo and in vitro experiments. RESULTS: hWJ-MSC attached to and proliferated on gelatin microbeads in 3D cultures reaching a maximum of 1.1-1.30×10(7) cells on 0.5 g of microbeads by days 8-14; in contrast, hWJ-MSC derived from 2D cultures reached a maximum of 6.5 -11.5×10(5) cells per well in a 24-well plate by days 6-10. hWJ-MSC derived by 3D culture incorporated significantly more EdU (P<0.05) and had a significantly higher proliferation index (P<0.05) than those derived from 2D culture. Immunofluorescence staining, real-time PCR, flow cytometry analysis, and multipotency assays showed that hWJ-MSC derived from 3D culture retained MSC surface markers and multipotency potential similar to 2D culture-derived cells. 3D culture-derived hWJ-MSC also retained the expression of core transcription factors at levels comparable to their 2D culture counterparts. Direct injection of hWJ-MSC derived from 3D or 2D cultures into animals exhibited similar efficacy in enhancing skin wound healing. CONCLUSIONS: Thus, hWJ-MSC can be expanded markedly in gelatin microbeads, while retaining MSC surface marker expression, multipotent differential potential, and expression of core transcription factors. These cells also efficiently enhanced skin wound healing in vivo, in a manner comparable to that of hWJ-MSC obtained from 2D culture.

Cyclosporine A increases hair follicle growth by suppressing apoptosis-inducing factor nuclear translocation: a new mechanism.

Cyclosporine A (CsA) enhances hair growth through caspase-dependent pathways by retarding anagen-to-catagen phase transition in the hair follicle growth cycle. Whether apoptosis-inducing factor (AIF), a protein that induces caspase-independent apoptosis, can regulate the hair follicle cycle in response to CsA is currently unclear. Here, we show that the pro-hair growth properties of CsA are in part due to blockage of AIF nuclear translocation. We first isolate hair follicles from murine dorsal skin. We then used Western blot, immunohistochemistry and immunofluorescence to evaluate the expression and localization of AIF in hair follicles. We also determined whether modulation of AIF was responsible for the effects of CsA at the anagen-to-catagen transition. AIF was expressed in hair follicles during the anagen, catagen and telogen phases. There was significant nuclear translocation of AIF as hair follicles transitioned from anagen to late catagen phase; this was inhibited by CsA, likely due to reduced cyclophilin A expression and attenuated AIF release from mitochondria. However, we note that AIF translocation was not completely eliminated, which likely explains why the transition to catagen phase was severely retarded by CsA, rather than being completely inhibited. We speculate that blockade of the AIF signalling pathway is a critical event required for CsA-dependent promotion of hair growth in mice. The study of AIF-related signalling pathways may provide insight into hair diseases and suggest potential novel therapeutic strategies.