Retinal safety and toxicity study of artesunate in vitro and in vivo.

Background: Artesunate (ART), a member of the artemisinin family, possesses multi-properties, including anti-inflammation, anti-oxidation, and anti-tumor. ART was recently reported to show anti-neovascularization effect on the cornea, iris, and retina. Compared to the expensive anti-VEGF treatment, this versatile, economical treatment option is attractive in the ophthalmic field. The safety and toxicity profile of ART intravitreal application are in utmost need. Methods: In this study, immortalized microglial (IMG) cells were treated with ART to determine the safe concentrations without inducing overt inflammatory reactions. Reverse transcription-polymerase chain reaction analysis was used to detect the cytokine expressions in IMG cells in response to ART stimulation. Various doses of ART were intravitreally injected into the right eyes of C57BL/6 mice. Retinal function was tested by electroretinogram, and retinal ganglion cell (RGC) survival was evaluated by counting Brn3a stained cells in flat-mounted retinas at 7 days after ART injection. Results: ART below 5µM was safe for IMG cells in vitro. Both 2.5 and 5 â€‹µM ART treatment increased IL-10 gene expression in IMG cells while not changing IL-1ß, IL-6, TNF-α, and Arg-1. In the in vivo study, intravitreal injection of ART below 100 â€‹µM did not cause deterioration in the retinal function and RGC survival of the mouse eyes, while 1 â€‹mM ART treatment significantly attenuated both the scotopic and photopic b-wave amplitudes and impaired RGC survival. In addition, treatment with ART of 25, 50, and 100 â€‹µM significantly decreased TNF-α gene expression while ART of 100 â€‹µM significantly increased IL-10 in the mouse retina. Conclusions: Intravitreal injection of 100 â€‹µM ART could downregulate TNF-α while upregulate IL-10 in the mouse retina without causing retinal functional deterioration and RGC loss. ART might be used as anti-inflammatory agent for retinal disorders.

Tumor Suppressor Role and Clinical Significance of the FEV Gene in Prostate Cancer.

Background: In our previous research, we developed a 32-gene risk index model that may be utilized as a robust prognostic method for predicting prostate cancer (PCa) recurrence after surgery. Among the 32 genes, the Fifth Ewing Variant (FEV) gene was one of the top downregulated genes in relapsed PCa. However, current understanding of the FEV gene and its involvement in PCa is limited. Methods: FEV mRNA expression was analyzed and correlated to clinical outcomes in PCa patients who underwent prostatectomy at the Massachusetts General Hospital. Specimens from tissue microarray (TMA) including 102 prostate cancer patients were analysis for the expression of FEV. Meanwhile, FEV expression profiles were also assessed in PCa cell lines and in BPH-1 prostate epithelial cells using western blotting and quantitative reverse transcription-PCR (qRT-PCR). Furthermore, we transfected LNCaP and PC-3 cells with either an empty vector or full-length FEV gene and performed in vitro cell functional assays. The part FEV plays in tumor xenograft growth was also assessed in vivo. Results: Of the 191 patients included in this study base on the DASL dataset, 77 (40.3%) and 24 (13.6%), respectively, developed prostate-specific antigen (PSA) relapse and metastasis postradical prostatectomy. Significant FEV downregulation was observed in PCa patients showing PSA failure and metastasis. The protein expression of FEV was significantly negatively correlated with the Gleason score and pathological stage in prostate cancer tissues. Similarly, FEV expression significantly decreased in all PCa cell lines relative to BPH-1 (all P < 0.05). Functional assays revealed that FEV expression markedly inhibited PCa cell growth, migration, and invasion, which in turn significantly repressed the growth of tumor xenografts in vivo. Conclusion: The results of this study suggest an association between downregulated FEV expression and PSA relapse in PCa patients. In addition, FEV may act as a tumor suppressor in PCa.

BHPF exposure impairs mouse and human decidualization.

Although BHPF has been widely used in plastic manufacturing as a substitute for BPA, current evidence suggests that BHPF also causes harmful effects on reproduction. However, effects of BHPF on mammalian early pregnancy are still poorly defined. This study aimed to explore the effects of BHPF on early pregnancy, especially decidualization and embryonic development in mice and human beings. The results showed that 50 and 100 mg/kg BHPF exposure reduced birth weight, and implantation site weight on the day 8 of pregnancy in mice. Because BHPF inhibits both embryo development and artificial decidualization in mice, suggesting that the detrimental effects of BHPF should be from its effects on embryo development and decidualization. Under in vitro decidualization, 10 µM BHPF inhibits decidualization and leads to disordered expression of Lamin B1 and collagen in mice. In addition, 10 µM BHPF also inhibits decidualization, and causes disordered expression of both collagen III and Lamin B1 under human in vitro decidualization. However, collagen III supplementation can rescue BHPF inhibition on decidualization. Further, our study demonstrates that BHPF impairs human decidualization through the HB-EGF/EGFR/STAT3/Collagen III pathway. Taken together these data suggest that exposure to BHPF impairs mouse and human decidualization during early pregnancy.

Clinical observations of vancomycin-loaded calcium phosphate cement in the 1-stage treatment of chronic osteomyelitis: a randomized trial.

BACKGROUND: This study sought to explore the clinical effects of vancomycin-loaded calcium phosphate cement (CPC) in the treatment of chronic osteomyelitis. METHODS: Ninety-eight patients with chronic osteomyelitis, who were treated at our Department from December 1st, 2014 to December 1st, 2015, were randomly allocated into the research group or the control group. Each group comprised 49 patients. Patients in the research group (Group A) received a 1-stage treatment of vancomycin-loaded CPC after debridement, while those in the control group (Group B) received an irrigation and drainage device that administered irrigated antibiotics. The treatment effects, recurrence rates, and safety outcomes of the two groups were observed. RESULTS: Patients in the two groups were followed up with for a period of 12 months. The control group without recurrence in one case. There was some systemic adverse postoperative effects and safety issues. The X-ray film showed that CPC filling is good and part of the degradation of osteogenesis. Some patients in the research group the CPC particles were absorbed, the bone defect area was healed, after One-year postoperatively. Further, the cure rate was high. CONCLUSIONS: One-stage vancomycin-loaded CPC implantation osteomyelitis lesions fill the die cavity, enable patients to continue to fight infection, induce bone defect osteogenesis, reduce the recurrence of chronic osteomyelitis, and are an effective method for treating chronic osteomyelitis. TRIAL REGISTRATION: Chinese Clinical Trial Registry website (ChiCTR2100044724).

Functional classification of prostate cancer­associated miRNAs through CRISPR/Cas9­mediated gene knockout.

The aim of the present study was to use the clustered regularly interspaced short palindromic repeats (CRISPR) and CRISPR­associated (Cas) 9­mediated gene knockout technology for the rapid classification of the differential function of micro (mi)RNAs screened using miRNA expression profiling by microarray. The rational design of single guide RNAs for the CRISPR/Cas9 system was verified to function in human LNCaP cells with rapid and efficient target gene editing. miRNA (miR)­205, miR­221, miR­222, miR­30c, miR­224, miR­455­3p, miR­23b and miR­505 were downregulated in patients with prostate cancer (PCa) and were experimentally validated to function as tumor suppressors in prostate cancer cells, affecting tumor proliferation, invasion and aerobic glycolysis. In addition, the data of the present study suggested that miR­663a and mfiR­1225­5p were upregulated in prostate cancer tissues and cell proliferation of miR­663a and miR­1225­5p knockout PCa cells was significantly lower compared with miR­NC cells. Furthermore, knockout of miR­1225­5p and miR­663a significantly decreased the lactate production in LNCaP cells in vitro. In conclusion, the present study offered a simple and efficient method for rapidly classifying miRNA function by applying CRISPR/Cas9 in LNCaP cells. The present study suggested, for the first time to the best of the authors' knowledge, that the aberrant expression of miR­663a and miR­1225­5p may be involved with the progression of prostate cancer, implying their potential as candidate markers for this type of cancer. However, the precise role of miR­663a and miR­1225­5p in accelerating the development of prostate cancer and promoting tumor progression remains to be elucidated.

Nucleolar stress regulates stromal-epithelial transition via NPM1 during decidualization.

Embryo implantation and decidualization are crucial steps during early pregnancy. We recently showed that nucleolar stress is involved in embryo implantation. This study was to explore whether nucleolar stress participates in mouse and human decidualization. Our data demonstrated that a low dose of actinomycin D (ActD) could induce nucleolar stress in stroma cells. Nucleolar stress promotes the stromal-epithelial transition during mouse in vitro decidualization through nucleophosmin1 (NPM1). Under nucleolar stress, Wnt family member 4 (Wnt4), a decidualization marker, is significantly increased, but decidua/trophoblast prolactin-related protein (Dtprp/Prl8a2) expression remains unchanged. For translational significance, we also examined the effects of nucleolar stress on human decidualization. Nucleolar stress stimulated by a low dose of ActD enhances human stromal-epithelial transition during human decidualization, but has no effects on the expression of insulin-like growth factor-binding protein 1 (IGFBP1). Our study indicates that nucleolar stress may promote only the mesenchymal-epithelial transition (MET), but not for all the molecular changes during decidualization.

ARNT-dependent CCR8 reprogrammed LDH isoform expression correlates with poor clinical outcomes of prostate cancer.

Lactate dehydrogenase isozyme (LDH) is a tetramer constituted of two isoforms, LDHA and LDHB, the expression of which is associated with cell metabolism and cancer progression. Our previous study reveals that CC-chemokine ligand-18 (CCL18) is involved in progression of prostate cancer (PCa).This study aims to investigate how CCL18 regulates LDH isoform expression, and therefore, contributes to PCa progression. The data revealed that the expression of LDHA was upregulated and LDHB was downregulated in PCa cells by CCL18 at both messenger RNA and protein levels. The depletion of CCR8 reduced the ability of CCL18 to promote the proliferation, migration, and lactate production of PCa cells. Depletion of a CCR8 regulated transcription factor, ARNT, significantly reduced the expression of LDHA. In addition, The Cancer Genome Atlas dataset analyses revealed a positive correlation between CCR8 and ARNT expression. Two dimension difference gel electrophoresis revealed that the LDHA/LDHB ratio was increased in the prostatic fluid of patients with PCa and PCa tissues. Furthermore, increased LDHA/LDHB ratio was associated with poor clinical outcomes of patients with PCa. Together, our results indicate that the CCR8 pathway programs LDH isoform expression in an ARNT dependent manner and that the ratio of LDHA/LDHB has the potential to serve as biomarkers for PCa diagnosis and prognosis.

Stavudine exposure results in developmental abnormalities by causing DNA damage, inhibiting cell proliferation and inducing apoptosis in mouse embryos.

Stavudine is an anti-AIDS drug widely used to prevent HIV transmission from pregnant mothers to the fetuses in underdeveloped countries for its low price. However, there is still a controversy on whether stavudine affects embryo development. In the current study, embryotoxicity of stavudine was evaluated using cultured mouse embryos with the concentrations: 5, 10, 15 µM and vehicle control. The data indicated that the effect of stavudine was dose-dependent at early neurogenesis. Stavudine exposure reduced somite numbers, yolk sac diameter, crown-rump length, and increased the rate of embryonic degeneration compared with the control. We chose the lowest but clearly toxic concentration: 5 µM to investigate the molecular mechanisms of the damage. At the molecular level, stavudine produced DNA damage, increased the levels of the phospho-CHK1 and cleaved-caspase-3, and decreased the expression level of proliferating cell nuclear antigen. These changes indicated that stavudine caused a coordinated DNA damage response, inhibited cell proliferation, and induced apoptosis in the embryos. Collectively these results suggest that stavudine exposure disturbs the embryonic development, and its use in pregnant mothers should be re-examined.

The study of multiple diagnosis models of human prostate cancer based on Taylor database by artificial neural networks.

BACKGROUND: Prostate cancer (PCa) is the most common malignancy seen in men and the second leading cause of cancer-related death in males. The incidence and mortality associated with PCa has been rapidly increasing in China recently. METHODS: Multiple diagnostic models of human PCa were developed based on Taylor database by combining the artificial neural networks (ANNs) to enhance the ability of PCa diagnosis. Genetic algorithm (GA) is used to select feature genes as numerical encoded parameters that reflect cancer, metastatic, or normal samples. Back propagation (BP) neural network and learning vector quantization (LVQ) neural network were used to build different Cancer/Normal, Primary/Metastatic, and Gleason Grade diagnostic models. RESULTS: The performance of these modeling approaches was evaluated by predictive accuracy (ACC) and area under the receiver operating characteristic curve (AUC). By observing the statistically significant parameters of the three training sets, our Cancer/Normal, Primary/Metastatic, and Gleason Grade models' with ACC and AUC can be drawn (97.33%, 0.9832), (99.17%, 0.9952), and (90.48%, 0.8742), respectively. CONCLUSION: These results indicated that our diagnostic models of human PCa based on Taylor database combining the feature gene expression profiling data and artificial intelligence algorithms might act as a powerful tool for diagnosing PCa. Gleason Grade diagnostic models were used as novel prognostic diagnosis models for biochemical recurrence-free survival and overall survival, which might be helpful in the prognostic diagnosis of PCa in patients.

Increasing of FKBP9 can predict poor prognosis in patients with prostate cancer.

BACKGROUND: FK506 binding protein 9 (FKBP9) has been reported and identified for a long time, but its relationship with cancer is rarely studied. For example, the role of FK506 binding protein 9 in prostate cancer (PCa) is still unclear. Therefore, we decided to detect the expression level of FKBP9 in PCa and explore its clinical significance. METHODS: The expression level of FKBP9 protein was detected by immunohistochemistry. In addition, it was demonstrated by high-throughput sequencing of mRNA levels in the TCGA (cancer genome atlas) dataset of 499 patients. Kaplan-meier analysis and Cox proportional hazard regression model were used to evaluate the relationship between FKBP9 expression and survival in prostate cancer patients. RESULTS: The expression of FKBP9 was localized in the cytoplasm, which in normal prostate tissues was obviously lower than that in PCa tissues (P = 0.001). High expression of FKBP9 was related with lymph node metastasis (P = 0.022) and distant metastasis (P = 0.028). Kaplan-Meier survival analysis revealed that the BCR-free survival of PCa patients with high FKBP9 level was significantly shortened (P=0.041). CONCLUSIONS: FKBP9 may be a cancer promoter that enhances PCa progression, and the level of FKBP9 may be used as an independent precursor of PCa patients.

Upregulation of Holliday junction recognition protein predicts poor prognosis and biochemical recurrence in patients with prostate cancer.

Abnormal expression of Holliday junction recognition protein (HJURP) in several types of tumor cells plays a vital role in the formation and progression of tumors. Few studies have investigated the role of HJURP in prostate cancer (PCa). The aim of this study was to analyze the expression levels of HJURP in PCa and to establish the association with clinicopathological data. Reverse transcription quantitative polymerase chain reaction and immunohistochemical analysis were used to detect the expression levels of HJURP in benign and PCa prostate tissues. The Taylor dataset was statistically analyzed to determine if HJURP expression levels were associated with PCa clinicopathological data. HJURP was overexpressed in PCa tissues compared with benign prostate tissues. Statistical analysis of the Taylor dataset indicated that upregulation of HJURP was significantly associated with positive prostate-specific antigen (PSA) levels (P=0.004), high Gleason score (P=0.005), advanced pathological stage (P=0.007), metastasis (P<0.001) and PSA failure (P<0.001). Higher HJURP mRNA expression levels were significantly associated with shorter biochemical recurrence (BCR)-free survival (P<0.001). To the best of our knowledge, this study is the first report of HJURP upregulation in PCa tissues. Upregulation of HJURP may predict BCR-free survival and HJURP may be an oncogene that impacts the prognosis of patients with PCa.

Nucleolar stress regulation of endometrial receptivity in mouse models and human cell lines.

Embryo implantation is essential to the successful establishment of pregnancy. A previous study has demonstrated that actinomycin D (ActD) could initiate the activation of mouse delayed implantation. However, the mechanism underlying this activation remains to be elucidated. A low dose of ActD is an inducer of nucleolar stress. This study was to examine whether nucleolar stress is involved in embryo implantation. We showed that nucleolar stress occurred when delayed implantation was activated by ActD in mice. ActD treatment also stimulated the Lif-STAT3 pathway. During early pregnancy, nucleolar stress was detected in the luminal epithelial cells during the receptive phase. Blastocyst-derived lactate could induce nucleolar stress in cultured luminal epithelial cells. The inhibition of nucleophosmin1 (NPM1), which was a marker of nucleolar stress, compromised uterine receptivity and decreased the implantation rates in pregnant mice. To translate these mouse data into humans, we examined nucleolar stress in human endometrium. Our data demonstrated that ActD-induced nucleolar stress had positive effects on the embryo attachment by upregulating IL32 expression in non-receptive epithelial cells rather than receptive epithelial cells. Our data should be the first to demonstrate that nucleolar stress is present during early pregnancy and is able to induce embryo implantation in both mice and humans.

Correction to: MicroRNA-30d promotes angiogenesis and tumor growth via MYPT1/c-JUN/VEGFA pathway and predicts aggressive outcome in prostate cancer.

After publication of the article [1], the author reported that this article contained some errors.

Overexpression of TPX2 is associated with progression and prognosis of prostate cancer.

Targeting protein for Xenopus kinesin-like protein 2 (TPX2) activates Aurora kinase A during mitosis and targets its activity to the mitotic spindle, serving an important role in mitosis. It has been associated with different types of cancer and is considered to promote tumor growth. The aim of the present study was to explore the role of TPX2 in diagnosing prostate cancer (PCa). It was identified that TPX2 expression in PCa tissues was increased compared with benign prostate tissues. Microarray analysis demonstrated that TPX2 was positively associated with the Gleason score, tumor-node-metastasis (TNM) stage, clinicopathological stage, metastasis, overall survival and biochemical relapse-free survival. In vitro studies revealed that the high expression of TPX2 in PCa cells improved proliferative, invasive and migratory abilities, and repressed apoptosis of the PCa cells, without affecting tolerance to docetaxel. The results suggested that TPX2 serves as a tumorigenesis-promoting gene in PCa, and a potential therapeutic target for patients with PCa.

Downregulation of ARID4A and ARID4B promote tumor progression and directly regulated by microRNA-30d in patient with prostate cancer.

AT-rich interaction domain 4A (ARID4A) and AT-rich interaction domain 4B (ARID4B), which are both the AT-rich interaction domain (ARID) family, have been reported to be oncogene or tumor suppressor gene in various human malignances, but there is no involvement about their functions in prostate cancer (PCa). Our previous study has reported that microRNA-30d (miR-30d) expression can predicted poor clinical prognosis in PCa, however, the underlying mechanisms of miR-30d have not been fully described. The aim of our study is to investigate the expression relevance between miR-30d and ARID4A or ARID4B, and examine the clinical significance and biological function of ARID4A and AIRD4B in PCa. In this study, both ARID4A and ARID4B were identified as the target genes of miR-30d. In addition, the mRNA expression of miR-30d in PCa tissues were significantly negative correlated with ARID4A (Pearson correlation coefficient = -0.313, P = 0.001) and ARID4B (Pearson correlation coefficient = -0.349, P < 0.001), while there was a positive correlation between ARID4A and ARID4B (Pearson correlation coefficient = 0.865, P < 0.001). Moreover, both ARID4A and ARID4B were significantly downregulated in PCa tissues with high Gleason scores (P = 0.005, P = 0.033), PSA failure (P = 0.012, P = 0.05) and short biochemical recurrent-free survival (P = 0.033, P = 0.031). Furthermore, the knockout expression of ARID4A and ARID4B promoted PCa cell proliferation, migration and invasion in vitro. In conclusion, our results indicated that ARID4A and ARID4B may serve as tumor suppressor in PCa progression, suggesting that they might be the potential therapeutic targets in prostate cancer.

Dual extra-retinal origins of microglia in the model of retinal microglia repopulation.

Elucidating the origin of microglia is crucial for understanding their functions and homeostasis. Previous study has indicated that Nestin-positive progenitor cells differentiate into microglia and replenish the brain after depleting most brain microglia. Microglia have also shown the capacity to repopulate the retina after eliminating all retinal microglia. However, the origin(s) of repopulated retinal microglia is/are unknown. In this study, we aim to investigate the origins of repopulated microglia in the retina. Interestingly, we find that repopulated retinal microglia are not derived from Nestin-positive progenitor cells. Instead, they have two origins: the center-emerging microglia are derived from residual microglia in the optic nerve and the periphery-emerging microglia are derived from macrophages in the ciliary body/iris. Therefore, we have for the first time identified the extra-retinal origins of microglia in the adult mammalian retina by using a model of microglial repopulation, which may shed light on the target exploration of therapeutic interventions for retinal degenerative disorders.

Repopulated microglia are solely derived from the proliferation of residual microglia after acute depletion.

Newborn microglia rapidly replenish the whole brain after selective elimination of most microglia (>99%) in adult mice. Previous studies reported that repopulated microglia were largely derived from microglial progenitor cells expressing nestin in the brain. However, the origin of these repopulated microglia has been hotly debated. In this study, we investigated the origin of repopulated microglia by a series of fate-mapping approaches. We first excluded the blood origin of repopulated microglia via parabiosis. With different transgenic mouse lines, we then demonstrated that all repopulated microglia were derived from the proliferation of the few surviving microglia (<1%). Despite a transient pattern of nestin expression in newly forming microglia, none of repopulated microglia were derived from nestin-positive non-microglial cells. In summary, we conclude that repopulated microglia are solely derived from residual microglia rather than de novo progenitors, suggesting the absence of microglial progenitor cells in the adult brain.

The high concentration of progesterone is harmful for endometrial receptivity and decidualization.

Progesterone is required for the establishment and maintenance of mammalian pregnancy and widely used for conservative treatment of luteal phase deficiency in clinics. However, there are limited solid evidences available for the optimal timing and dose of progesterone therapy, especially for the possible adverse effects on implantation and decidualization when progesterone is administrated empirically. In our study, mouse models were used to examine effects of excess progesterone on embryo implantation and decidualization. Our data indicate that excess progesterone is not only harmful for mouse implantation, but also impairs mouse decidualization. In excess progesterone-treated mice, the impaired LIF/STAT3 pathway and dysregulated endoplasmic reticulum stress may lead to the inhibition of embryo implantation and decidualization. It is possible that the decrease in birth weight of excess progesterone-treated mice is due to a compromised embryo implantation and decidualization. Furthermore, excess progesterone compromises in vitro decidualization of human endometrial stromal cells.