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Background There has been a longstanding debate about the role of folate in the etiology of orofacial clefts (OFCs). Studies of different measures of nutritional intake or folate status have been done to investigate the possible role of folate in the prevention of OFC. Only one knowledge synthesis has attempted to bring together different types of evidence. The aim of the present work was to update it. Methods Evidence for associations between OFC and dietary folate, supplement use, folic acid fortification, biomarkers of folate status, and variants of MTHFR (C677T and A1298C) were included. Potentially eligible articles were systematically identified from PubMed, Medline, Embase, and Web of Science (2007-2020) and combined using random-effects meta-analysis when appropriate. Quality assessments were conducted using the Newcastle-Ottawa scale and Cochrane's risk of bias tool. Results Sixty-four studies published since the previous knowledge synthesis were identified, with eight of these identified through a supplementary search from October, 2018 to August, 2020. There was an inverse association between folic acid-containing supplement use before or during pregnancy and cleft lip with or without cleft palate (CL/P) (OR 0.60, 95% CI 0.51-0.69), with considerable between-study heterogeneity. The prevalence of CL/P showed a small decline post-folic acid fortification in seven studies (OR 0.94, 95% CI 0.86-1.02). No association was found between OFC and genetic markers of folate status. The coronavirus-19 pandemic has threatened food availability globally and therefore there is a need to maintain and even enhance surveillance concerning maternal intake of folate and related vitamins. Conclusions The risk of non-syndromic OFC was reduced among pregnant women with folic acid-containing supplements during the etiologically relevant period. However, high heterogeneity between included studies, incomplete reporting of population characteristics and variation in timing of exposure and supplement types mean that conclusions should be drawn with caution.

In advanced prostate cancer, CREB (cAMP-responsive element-binding protein) binding protein (CBP) and its homolog EP300 are highly expressed; targeting the bromodomain of CBP is a new strategy for the treatment of prostate cancer. In the current study we identified Y08197, a novel 1-(indolizin-3-yl) ethanone derivative, as a selective inhibitor of CBP/EP300 bromodomain and explored its antitumor activity against prostate cancer cell lines in vitro. In the AlphaScreen assay, we demonstrated that Y08197 dose-dependently inhibited the CBP bromodomain with an IC(50) value at 100.67 ± 3.30 nM. Y08197 also exhibited high selectivity for CBP/EP300 over other bromodomain-containing proteins. In LNCaP, 22Rv1 and VCaP prostate cancer cells, treatment with Y08197 (1, 5 μM) strongly affected downstream signaling transduction, thus markedly inhibiting the expression of androgen receptor (AR)-regulated genes PSA, KLK2, TMPRSS2, and oncogenes C-MYC and ERG. Notably, Y08197 potently inhibited cell growth in several AR-positive prostate cancer cell lines including LNCaP, 22Rv1, VCaP, and C4-2B. In 22Rv1 prostate cancer cells, treatment with Y08197 (1, 4, 16 μM) dose-dependently induced G(0)/G(1) phase arrest and apoptosis. Furthermore, treatment with Y08197 (5 μM) significantly decreased ERG-induced invasive capacity of 22Rv1 prostate cancer cells detected in wound-healing assay and cell migration assay. Taken together, CBP/EP300 inhibitor Y08197 represents a promising lead compound for development as new therapeutics for the treatment of castration-resistant prostate cancer.

Glioblastoma (GBM) continues to be the most aggressive cancer of the brain. The dismal prognosis is largely attributed to the microenvironment surrounding tumor cells. Astrocytes, the main component of the GBM microenvironment, play several fundamental physiological roles in the central nervous system. During the development of GBM, tumor-associated astrocytes (TAAs) directly contact GBM cells, which activate astrocytes to form reactive astrocytes, facilitating tumor progression, proliferation and migration through multiple well-understood signaling pathways. Notably, TAAs also influence GBM cell behaviors via suppressing immune responses and enhancing the chemoradiotherapy resistance of tumor cells. These new activities are closely linked with the treatment and prognosis of GBM. In this review, we discuss recent advances regarding new functions of reactive astrocytes, including TAA-cancer cell interactions, mechanisms involved in immunosuppressive regulation, and chemoradiotherapy resistance. It is expected that these updated experimental or clinical studies of TAAs may provide a promising approach for GBM treatment in the near future.

The development of induced pluripotent stem cells (iPS cells) has raised the prospect of patient-specific treatments for various diseases. Theoretically, iPS cell technology avoids the limitations of human embryonic stem cells (ES cells), including poor establishment, ethical issues, and immune rejection of allogeneic transplantation. However, the immunogenicity of iPS cells has attracted the attention of researchers, and it remains unclear whether iPS cells and their derivatives will be recognized as a patient's own cells. Even though iPS-derived functional cells have been used in the treatment of some diseases, the process of somatic cell reprogramming and iPS cell differentiation is time-consuming, making it difficult to use iPS cells in acute illness or injury. In recent years, it has been suggested that iPS cells may be used as allografts by establishing an iPS cell bank and HLA matching, providing a novel strategy for the clinical application of iPS cells. This article provides a concise overview of iPS cell immunogenicity, and summarizes published data regarding the application of iPS cells in both autologous and allogeneic transplantation in order to help develop more reliable biotechnical strategies utilizing iPS cells.

The CREB (cAMP responsive element binding protein) binding protein (CBP) and its homolog EP300 have emerged as new therapeutic targets for the treatment of cancer and inflammatory diseases. Here we report the identification, optimization and evaluation of 1-(1H-indol-1-yl)ethanone derivatives as CBP/EP300 inhibitors starting from fragment-based virtual screening (FBVS). A cocrystal structure of the inhibitor (22e) in complex with CBP provides a solid structural basis for further optimization. The most potent compound 32h binds to the CBP bromodomain and has an IC50 value of 0.037 μM in the AlphaScreen assay which was 2 times more potent than the reported CBP bromodomain inhibitor SGC-CBP30 in our hands. 32h also exhibit high selectivity for CBP/EP300 over other bromodomain-containing proteins. Notably, the ester derivative (29h) of compound 32h markedly inhibits cell growth in several prostate cancer cell lines including LNCaP, 22Rv1 and LNCaP derived C4-2B. Compound 29h suppresses the mRNA expression of full length AR (AR-FL), AR target genes and other oncogene in LNCaP cells. 29h also reduces the expression of PSA, the biomarker of prostate cancer. CBP/EP300 inhibitor 29h represents a promising lead compound for the development of new therapeutics for the treatment of castration-resistant prostate cancer.

Browning in adipose tissues, which can be affected by diet, may mitigate the detrimental effects of adiposity and improve longer-term metabolic health. Here, browning-inducing effects of long-chain polyunsaturated fatty acids, e.g., arachidonic acid (ARA)/docosahexaenoic acid (DHA) and extensively hydrolyzed casein (eHC) were investigated in uncoupling protein 1 (Ucp-1) reporter mice. To address the overall functionality, their potential role in supporting a healthy metabolic profile under obesogenic dietary challenges later in life was evaluated. At weaning Ucp1+/LUC reporter mice were fed a control low fat diet (LFD) with or without ARA + DHA, eHC or eHC + ARA + DHA for 8 weeks until week 12 after which interventions continued for another 12 weeks under a high-fat diet (HFD) challenge. Serology (metabolic responses and inflammation) and in vivo and ex vivo luciferase activity were determined; in the meantime browning-related proteins UCP-1 and the genes peroxisome proliferator-activated receptor gamma coactivator 1-alpha (PGC1α), PR domain containing 16 (PRDM16) and Ucp-1 were examined. ARA + DHA, eHC or their combination reduced body weight gain and adipose tissue weight compared to the HFD mice. The interventions induced Ucp-1 expression in adipose tissues prior to and during the HFD exposure. Ucp-1 induction was accompanied by higher PGC1a and PRDM16 expression. Glucose tolerance and insulin sensitivity were improved coinciding with lower serum cholesterol, triglycerides, free fatty acids, insulin, leptin, resistin, fibroblast growth factor 21, alanine aminotransferase, aspartate aminotransferase and higher adiponectin than the HFD group. HFD-associated increased systemic (IL-1β and TNF-α) and adipose tissue inflammation (F4/80, IL-1β, TNF-α, IL-6) was reduced. Studies in a Ucp-1 reporter mouse model revealed that early intervention with ARA/DHA and eHC improves metabolic flexibility and attenuates obesity during HFD challenge later in life. Increased browning is suggested as, at least, part of the underlying mechanism.

Contiguous microdeletions of the Norrie disease pseudoglioma (NDP) region on chromosome Xp11.3 have been widely confirmed as contributing to the typical clinical features of Norrie disease (ND). However, the precise relation between genotype and phenotype could vary. The contiguous deletion of NDP and its neighbouring genes, MAOA/B and EFHC2, reportedly leads to syndromic clinical features such as microcephaly, intellectual disability, and epilepsy. Here we report a novel contiguous microdeletion of the NDP region containing the MAOB and EFHC2 genes, which causes eye defects but no cognitive disability. We detected a deletion of 494.6 kb at Xp11.3 in both the proband and carrier mother. This deletion was then used as the molecular marker in prenatal diagnosis for two subsequent pregnancies. The deletion was absent in one of the foetuses, who remain without any abnormalities at 2 years of age. The proband shows the typical ocular clinical features of ND including bilateral retinal detachment, microphthalmia, atrophic irides, corneal opacification, and cataracts, but no symptoms of microcephaly, intellectual disability, and epilepsy. This familial study demonstrates that a deficiency in one of two MAO genes may not lead to psychomotor delay, and deletion of EFHC2 may not cause epilepsy. Our observations provide new information on the genotype–phenotype relations of $${\textit{MAOA/B}}$$ and EFHC2 genes involved in the contiguous deletions of ND.

Cong Hao,1–3 Xuewen Wu,1–3 Ruoyu Zhou,1–3 Hao Zhang,1,2 Yulai Zhou,1,2 Xinxing Wang,1,2 Yong Feng,1–3 Lingyun Mei,1–3 Chufeng He,1–3 Xinzhang Cai,1–3 Lisha Wu1–3 1Department of Otolaryngology Head and Neck Surgery, Xiangya Hospital Central South University, Changsha 410008, Hunan, People’s Republic of China; 2Hunan Province Key Laboratory of Otolaryngology Critical Diseases, Changsha 410008, Hunan, People’s Republic of China; 3National Clinical Research Center for Geriatric Disorders, Xiangya Hospital, Central South University, Changsha 410008, Hunan, People’s Republic of ChinaCorrespondence: Lisha WuDepartment of Otolaryngology Head and Neck Surgery, Xiangya Hospital Central South University, Changsha 410008, Hunan, People’s Republic of ChinaEmail ENTLisaWu@163.comBackground: p66Shc, a Src homologue and collagen homologue (Shc) adaptor protein, mediates oxidative stress signaling. The p66Shc-null mice have increased lifespan and enhanced resistance to oxidative stress. Studies have also indicated its potential role in inner ear aging, which can lead to deafness.Objective: The aim of this study was to determine the effects of p66Shc down-regulation on the marginal cells (MCs) of the inner ear stria vascularis.Methods: Primary MCs were isolated from neonatal rats and treated with glucose oxidase to induce oxidative stress. The cells were transduced with adenovirus expressing siRNA, and the knockdown was verified by Western blotting. The reactive oxygen species (ROS) levels and apoptosis were analyzed using the DCFH-DA probe and Annexin-V/7-AAD staining respectively. The ultrastructure of the differentially-treated cells was examined by transmission electron microscopy (TEM).Results: The in vitro oxidative stress model was established successfully in rat MCs. Knockdown of p66Shc alleviated the high ROS levels and apoptosis in the glucose oxidase-treated cells. In addition, glucose oxidase significantly increased the number of peroxisomes in the MCs, which was decreased by p66Shc inhibition.Conclusion: Oxidative stress increases p66Shc levels in the marginal cells of the inner ear, which aggravates ROS production and cellular injury. Blocking p66Shc expression can effectively reduce oxidative stress and protect the MCs.Keywords: p66Shc, marginal cells of stria vascularis, oxidative stress, peroxisome &nbsp

Multiple myeloma (MM) is an incurable hematological malignancy, although bortezomib has markedly improved its outcomes. Growing clinical evidence indicates that enhancing induced natural killer (NK) or γδ T cells for infusion is useful in the treatment of MM. However, whether combination treatment with bortezomib and induced NK and γδ T cells further improves outcomes in MM, and how the treatments should be combined, remain unclear. Herein, we found that low-dose bortezomib did not suppress the viability of induced NK and γδ T cells, but did induce MM cell apoptosis. Importantly, low-dose bortezomib increased the expression of NKG2D and DNAM-1 ligands on MM cells, which sensitized the multiple myeloma cells to lysis by induced NK and γδ T cells. Our results suggested that combination treatment with low-dose bortezomib and induced NK or γδ T cells had a synergistic cytotoxic effect on MM cells. This study provided a proof of principle for the design of future trials and investigation of this combination therapeutic strategy for MM treatment.

Aims The aim of this study was to improve a method that induce cartilage differentiation of human embryoid stem cells (hESCs) in vitro, and test the effect of in vivo environments on the further maturation of hESCs derived cells. Main methods Embryoid bodies (EBs) formed from hESCs, with serum-free KSR-based medium and mesodermal specification related factors, CHIR, and Noggin for first 8 days. Then cells were digested and cultured as micropellets in serum-free KSR-based chondrogenic medium that was supplemented with PDGF-BB, TGF β3, BMP4 in sequence for 24 days. The morphology, FACS, histological staining as well as the expression of chondrogenic specific genes were detected in each stage, and further in vivo experiments, cell injections and tissue transplantations, further verified the formation of chondrocytes. Key findings We were able to obtain chondrocyte/cartilage from hESCs using serum-free KSR-based conditioned medium. qPCR analysis showed that expression of the chondroprogenitor genes and the chondrocyte/cartilage matrix genes. Morphology analysis demonstrated we got PG + COL2 + COL1-particles. It indicated we obtained hyaline cartilage-like particles. 32-Day differential cells were injected subcutaneous. Staining results showed grafts developed further mature in vivo. But when transplanted in subrenal capsule, their effect was not good as in subcutaneous. Microenvironment might affect the cartilage formation. Significance The results of this study provide an absolute serum-free and efficient approach for generation of hESC-derived chondrocytes, and cells will become further maturation in vivo. It provides evidence and technology for the hypothesis that hESCs may be a promising therapy for the treatment of cartilage disease.

Transcription factor nuclear factor-erythroid 2-like 2 (NRF2) mainly regulates cellular antioxidant response, redox homeostasis and metabolic balance. Our previous study illustrated the translational significance of NRF2-mediated transcriptional repression, and the transcription of FOCAD gene might be negatively regulated by NRF2. However, the detailed mechanism and the related significance remain unclear. In this study, we mainly explored the effect of NRF2-FOCAD signaling pathway on ferroptosis regulation in human non-small-cell lung carcinoma (NSCLC) model. Our results confirmed the negative regulation relationship between NRF2 and FOCAD, which was dependent on NRF2-Replication Protein A1 (RPA1)-Antioxidant Response Elements (ARE) complex. In addition, FOCAD promoted the activity of focal adhesion kinase (FAK), which further enhanced the sensitivity of NSCLC cells to cysteine deprivation-induced ferroptosis via promoting the tricarboxylic acid (TCA) cycle and the activity of Complex I in mitochondrial electron transport chain (ETC). However, FOCAD didn't affect GPX4 inhibition-induced ferroptosis. Moreover, the treatment with the combination of NRF2 inhibitor (brusatol) and erastin showed better therapeutic action against NSCLC in vitro and in vivo than single treatment, and the improved therapeutic function partially depended on the activation of FOCAD-FAK signal. Taken together, our study indicates the close association of NRF2-FOCAD-FAK signaling pathway with cysteine deprivation-induced ferroptosis, and elucidates a novel insight into the ferroptosis-based therapeutic approach for the patients with NSCLC.

Obesity is caused by energy imbalance and accompanied by adipocyte hypertrophy and hyperplasia. Therefore, both enhancement of adipocyte energy expenditure and inhibition of adipogenesis are viable ways to combat obesity. Using the Ucp1-2A-luciferase reporter animal model previously reported by us as a screening platform, a chemical compound Linifanib was identified as a potent inducer of UCP1 expression in primary inguinal adipocytes in vitro and in vivo. Signal pathway analyses showed that Linifanib promoted adipocyte browning by attenuating STAT3 phosphorylation. The effects of Linifanib on adipocyte browning were blocked by the compound, SD19, which activates the STAT3 signaling cascade. Linifanib also inhibited adipocyte differentiation, by blocking mitotic clonal expansion, which could be rescued by STAT3 activator. Taken together, our results indicate that Linifanib might serve as a potential drug for the treatment of obesity.

[Image: see text] Prostate cancer is a commonly diagnosed cancer and a leading cause of cancer-related deaths. The bromodomain and extra terminal domain (BET) family proteins have emerged as potential therapeutic targets for the treatment of castration-resistant prostate cancer. A series of 2,2-dimethyl-2H-benzo[b][1,4]oxazin-3(4H)-one derivatives were designed and synthesized as selective bromodomain containing protein 4 (BRD4) inhibitors. The compounds potently inhibit BRD4(1) with nanomolar IC(50) values and exhibit high selectivity over most non-BET subfamily members. One of the representative compounds 36 (Y08060) effectively suppresses cell growth, colony formation, and expression of androgen receptor (AR), AR regulated genes, and MYC in prostate cancer cell lines. In in vivo studies, 36 demonstrates a good PK profile with high oral bioavailability (61.54%) and is a promising lead compound for further prostate cancer drug development.

As a member of the phenothiazine family, thioridazine (THIO) is a potent anti-anxiety and anti-psychotic drug. Recent studies have reported that THIO could suppress the growth of several types of cancer cells. However, the effect of THIO on colorectal cancer stem cells (CSCs) has not been investigated. In the present study, we examined the effect of THIO on viability of CSCs isolated from the human colon cancer cell line HCT116 and its colony-formation ability, along with its stem cell-specific gene expression. The CSCs, EpCAM+ and CD44+ subpopulations from HCT116 cells were isolated using immunomagnetic beads. After incubation with several concentrations of THIO, we evaluated the proliferation and invasion ability of colon CSCs, as well as cell apoptosis. We found that THIO significantly suppressed the proliferation and invasion of colon CSCs and induced cell apoptosis in a concentration-dependent manner. The expression of some apoptosis genes (Bax and caspase-3) was upregulated after treatment with THIO, while that of the anti-apoptosis gene Bcl-2 was downregulated. Moreover, the CSC mitochondrial membrane potential was downregulated. Overall, this study showed that THIO inhibits the proliferation of CSCs derived from the HCT116 cell line through induction of apoptosis, and thus, could be a promising agent for the treatment of colon cancer and worthy of exploring in prospective clinical studies.

The therapeutic action of umbilical cord-derived mesenchymal stem cells (UC-MSCs) against acute kidney injury (AKI) has been demonstrated by several groups. However, how to further enhance the renoprotective effect of UC-MSCs and improve the therapy effect, are still unclear. In this study, we mainly investigated whether insulin-like growth factor-1 (IGF-1)-modified UC-MSCs hold an enhanced protective effect on gentamicin-induced AKI in vivo. Our results indicated that the IGF-1 overexpression could enhance the therapeutic action of human UC-MSCs and the AKI rats treated with IGF-1-overexpressed UC-MSCs (UC-MSCs-IGF-1) showed better recovery of biochemical variables in serum or urine associated with renal function, histological injury and renal apoptosis, compared with AKI rats treated with normal UC-MSCs. RNA microarray analysis indicated that some key genes in the signal pathways associated with anti-oxidation, anti-inflammatory and cell migratory capacity were up-regulated in UC-MSCs-IGF-1 and the results were further confirmed with qPCR. Furthermore, a series of detection in vitro and in vivo indicated that the UC-MSCs-IGF-1 hold better anti-oxidation, anti-inflammatory and cell migratory capacity for IGF-1 overexpression. Thus, our study indicated that enhancement of UC-MSCs bioactivities with IGF-1 overexpression could increase the UC-MSCs therapeutic potential and further developed a new therapeutic strategy for the treatment of AKI.

Objective. To explore the effects of low concentration of exogenous fetuin-A intervention on TGF-β1 induced LX2 cells through detection of the expression of mRNA and protein of Smad2, Smad3, and Smad7. Methods. MTT assay was used to detect the LX2 cells proliferation and the regression equation calculating software was applied to determine IC50 of fetuin-A. RT-PCR was used to determine the relative content of Smad2, Smad3, and Smad7 mRNA in LX2 cells. Western blot was used to detect the LX2 cells relative content of Smad2, Smad3, Smad7 protein expression, respectively. Results. The analysis from RT-PCR and western blot showed that when compared with the other groups TGF-β1 + fetuin-A group increased the expression of Smad2 and Smad3 while decreased the expression of Smad7 (P<0.05). Conclusion. Fetuin-A may improve the excessive activation of hepatic stellate cells which is caused by an enhanced positive regulation of Smad2 and Smad3 protein and the deficiency in negative regulation of Smad7 protein. This is through inhibiting the expression of Smad2 and Smad3 gene and promoting the expression of Smad7 gene. As a result, the development of liver fibrosis will be reduced.