Phosphorylation of IDH1 Facilitates Progestin Resistance in Endometrial Cancer.

The progestin regimen is one of the main therapeutic strategies for women with endometrial cancer who undergo conservative management. Although many patients respond well to initial therapy, progestin-refractory disease inevitably emerges, and the molecular basis underlying progestin resistance has not been comprehensively elucidated. Herein, they demonstrated progestin results in p38-dependent IDH1 Thr 77 phosphorylation (pT77-IDH1). pT77-IDH1 translocates into the nucleus and is recruited to chromatin through its interaction with OCT6. IDH1-produced α-ketoglutarate (αKG) then facilitates the activity of OCT6 to promote focal adhesion related target gene transcription to confer progestin resistance. Pharmacological inhibition of p38 or focal adhesion signaling sensitizes endometrial cancer cells to progestin in vivo. The study reveals p38-dependent pT77-IDH1 as a key mediator of progestin resistance and a promising target for improving the efficacy of progestin therapy.

TSLP enhances progestin response in endometrial cancer via androgen receptor signal pathway.

BACKGROUND: The enriched proteins within in vitro fertilisation (IVF)-generated human embryonic microenvironment could reverse progestin resistance in endometrial cancer (EC). METHODS: The expression of thymic stromal lymphopoietin (TSLP) in EC was evaluated by immunoblot and IHC analysis. Transcriptome sequencing screened out the downstream pathway regulated by TSLP. The role of TSLP, androgen receptor (AR) and KANK1 in regulating the sensitivity of EC to progestin was verified through a series of in vitro and in vivo experiments. RESULTS: TSLP facilitates the formation of a BMP4/BMP7 heterodimer, resulting in activation of Smad5, augmenting AR signalling. AR in turn sensitises EC cells to progestin via KANK1. Downregulation of TSLP, loss of AR and KANK1 in EC patients are associated with tumour malignant progress. Moreover, exogenous TSLP could rescue the anti-tumour effect of progestin on mouse in vivo xenograft tumour. CONCLUSIONS: Our findings suggest that TSLP enhances the sensitivity of EC to progestin through the BMP4/Smad5/AR/KANK1 axis, and provide a link between embryo development and cancer progress, paving the way for the establishment of novel strategy overcoming progestin resistance using embryo original factors.

Partial substitution of whey protein concentrate by zein in high-protein nutrition bars: An effective method to reduce hardening during storage.

Whey protein concentrate-based high-protein nutrition bars (WPC-based HPN bars) are prone to hardening during storage, which limits their shelf life. In this study, zein was introduced to partially substitute WPC in the WPC-based HPN bars. The result of storage experiment revealed that the hardening of WPC-based HPN bars was significantly reduced with increasing zein content from 0% to 20% (mass ratio, zein:WPC-based HPN bar). Subsequently, the possible anti-hardening mechanism of zein substitution was studied in detail by determining the change in microstructure, patterns, free sulfhydryl group, color, free amino group, and Fourier transform infrared spectra of WPC-based HPN bars during storage. The results showed that zein substitution significantly blocked protein aggregation by inhibiting cross-linking, the Maillard reaction, and protein secondary structure transformation from α-helix to ß-sheet, which reduced the hardening of WPC-based HPN bars. This work provides insight into the potential utilization of zein substitution to improve the quality and shelf life of WPC-based HPN bars. PRACTICAL APPLICATION: In the preparation of whey protein concentrate-based high-protein nutrition bars, the introduction of zein to partially replace WPC can effectively reduce the hardening of WPC-based HPN bars during storage by preventing protein aggregation between WPC macromolecules. Therefore, zein could act as an agent to reduce the hardening of WPC-based HPN bars.

Diverse Effects of Amino Acids on Monascus Pigments Biosynthesis in Monascus purpureus.

Amino acids could act as nitrogen sources, amido group donors, or bioactive molecules in fungi fermentation, and consequently, play important roles in Monascus pigments (MPs) biosynthesis. But the understanding of the effects of various amino acids on MPs biosynthesis is still incomprehensive. In this work, 20 free amino acids were added to the fermentation medium to evaluate their effects on MPs biosynthesis in Monascus purpureus RP2. Six amino acids, namely, histidine (HIS), lysine (LYS), tyrosine (TYR), phenylalanine (PHE), methionine (MET), and cysteine (CYS), were selected as the valuable ones as they exerted significant effects on the production yield and even on the biosynthesis metabolic curves of MPs. Moreover, the dose-dependent and synergistic effects of valuable amino acids on MPs biosynthesis were observed by tests of serial concentrations and different combinations. In addition, it revealed that HIS and MET were the prominent amino acids with dominant and universal influences on MPs biosynthesis. The analog compounds of HIS (amitrole) and MET [calcium 2-hydroxy-4-(methylthio)] were added to the fermentation medium, and the results further confirmed the extraordinary effects of HIS and MET and their analogs on MPs biosynthesis. Furthermore, the gene transcription profile indicated that a differential expression pattern was observed in the polyketide synthase (PKS) cluster responsible for MPs biosynthesis in response to HIS and MET, revealing that they could oppositely regulate MPs biosynthesis in different ways. These findings would benefit the understanding of MPs biosynthesis regulation mechanism in M. purpureus and contribute to the industrial production of MPs by fermentation.

Brusatol sensitizes endometrial hyperplasia and cancer to progestin by suppressing NRF2-TET1-AKR1C1-mediated progestin metabolism.

Progestin resistance is the main obstacle for the conservative therapy to maintain fertility in women with endometrial cancer. Brusatol was identified as an inhibitor of the NRF2 pathway; however, its impact on progestin resistance and the underlying mechanism remains unclear. Here, we found that brusatol sensitized endometrial cancer to progestin by suppressing NRF2-TET1-AKR1C1-mediated progestin metabolism. Brusatol transcriptionally suppressed AKR1C1 via modifying the hydroxymethylation status in its promoter region through TET1 inhibition. Suppression of AKR1C1 by brusatol resulted in decreased progesterone catabolism and maintained potent progesterone to inhibit endometrial cancer growth. This inhibition pattern has also been found in the established xenograft mouse and organoid models. Aberrant overexpression of AKR1C1 was found in paired endometrial hyperplasia and cancer samples from the same individuals with progestin resistance, whereas attenuated or loss of AKR1C1 was observed in post-treatment samples with well progestin response as compared with paired pre-treatment tissues. Our findings suggest that AKR1C1 expression pattern may serve as an important biomarker of progestin resistance in endometrial cancer.

Methionine and S-Adenosylmethionine Regulate Monascus Pigments Biosynthesis in Monascus purpureus.

Amino acid metabolism could exert regulatory effects on Monascus pigments (MPs) biosynthesis. In this work, MPs biosynthesis regulated by methionine and S-adenosylmethionine (SAM) was investigated in Monascus purpureus RP2. The results indicated that the addition of methionine in fermentation significantly reduced MPs production by 60-70%, and it induced a higher expression of SAM synthetase Mon2A2272 and consequently led to SAM accumulation. However, the addition of SAM in fermentation promoted MPs production by a maximum of 35%, while over-expression of the gene Mon2A2272 led to a decrease in MPs yield, suggesting that SAM synthetase and SAM were likely to play different regulatory roles in MPs biosynthesis. Furthermore, the gene transcription profile indicated that SAM synthetase expression led to a higher expression of the transcriptional regulatory protein of the MPs biosynthesis gene cluster, while the addition of SAM gave rise to a higher expression of MPs biosynthesis activator and the global regulator LaeA, which probably accounted for changes in MPs production and the mycelium colony morphology of M. purpureus RP2 triggered by methionine and SAM. This work proposed a possible regulation mechanism of MPs biosynthesis by SAM metabolism from methionine. The findings provided a new perspective for a deep understanding of MPs biosynthesis regulation in M. purpureus.

The CD8α-PILRα interaction maintains CD8+ T cell quiescence.

T cell quiescence is essential for maintaining a broad repertoire against a large pool of diverse antigens from microbes and tumors, but the underlying molecular mechanisms remain largely unknown. We show here that CD8α is critical for the maintenance of CD8+ T cells in a physiologically quiescent state in peripheral lymphoid organs. Upon inducible deletion of CD8α, both naïve and memory CD8+ T cells spontaneously acquired activation phenotypes and subsequently died without exposure to specific antigens. PILRα was identified as a ligand for CD8α in both mice and humans, and disruption of this interaction was able to break CD8+ T cell quiescence. Thus, peripheral T cell pool size is actively maintained by the CD8α-PILRα interaction in the absence of antigen exposure.

B7-H3 specific T cells with chimeric antigen receptor and decoy PD-1 receptors eradicate established solid human tumors in mouse models.

The application of chimeric antigen receptor (CAR)-T cell therapy in patients with advanced solid tumors remains a significant challenge. Simultaneously targeting antigen and the solid tumor microenvironment are two major factors that greatly impact CAR-T cell therapy outcomes. In this study, we engineered CAR-T cells to specifically target B7-H3, a protein commonly found in solid human tumors, using a single-chain variable fragment (scFv) derived from an anti-B7-H3 monoclonal antibody. We tested the antitumor activity of B7-H3 CAR-T cells in mouse models with solid human tumors and determined that B7-H3 CAR-T cells exhibited potent antitumor activity against B7-H3+ tumor cells in vitro and in vivo. In addition, PD-1 decoy receptors were engineered to include extracellular PD-1 fused to the intracellular stimulatory domain of either CD28 or IL-7 receptor, respectively, which were then introduced into B7-H3 CAR-T cells. As a result, these newly modified, superior CAR-T cells exhibited more persistent antitumor activity in B7-H3+/B7-H1+ tumors in vivo. Our findings indicate that B7-H3 specific CAR-T cells have the potential to treat multiple types of advanced solid tumors.

Low-fat yogurt alleviates the pro-inflammatory cytokine IL-1ß-induced intestinal epithelial barrier dysfunction.

Yogurt is a source of bioactive compounds and probiotic microorganisms that modify immunity and metabolism to benefit human health beyond nutrition. In this study, we examined the capacity of yogurt to prevent epithelial barrier disruption in vitro. Different preparations of yogurt were added apically to Caco-2 monolayers before IL-1ß exposure. Dulbecco's modified Eagle medium containing 25% (vol/vol) low-fat yogurt prevented cytokine-induced transepithelial resistance reduction and increases to paracellular permeability measured with fluorescein isothiocyanate-dextran (4 kDa), whereas nonfat yogurt was unable to decrease paracellular permeability to fluorescein isothiocyanate-dextran. Moreover, the concentration of IL-8 in low-fat-yogurt-treated inflamed cells was decreased to 252.40 ± 27.24 pg/mL, which was lower than that of untreated, inflamed cells (407.20 ± 50.05 pg/mL), further indicating the anti-inflammatory roles of low-fat yogurt. The low-fat yogurt was able to downregulate the transcription of myosin light-chain kinase (MLCK) gene, but upregulate the expression of tight junction protein ZO-1 (TJP1). These findings indicate that low-fat yogurt can maintain intestinal barrier integrity better than nonfat yogurt after pro-inflammatory cytokine exposure.

B7-DC (PD-L2) costimulation of CD4+ T-helper 1 response via RGMb.

The role of B7-DC in T-cell responses remains controversial because both coinhibitory and costimulatory functions have been reported in various experimental systems in vitro and in vivo. In addition to interacting with the coinhibitory receptor PD-1, B7-DC has also been shown to bind repulsive guidance molecule b (RGMb). The functional consequences of the B7-DC/RGMb interaction, however, remain unclear. More than a decade ago, we reported that replacement of a murine B7-DC mutant lysine with serine (K113S) at positive 113 resulted in a loss of binding capacity to PD-1. Nevertheless, K113S remained costimulatory for T cells in vitro, implicating a dual functionality for B7-DC in T-cell responses. Here we show that recombinant K113S protein interacts with RGMb with a similar affinity to wild-type B7-DC. More importantly, K113S costimulates CD4+ T-cell responses via RGMb and promotes Th1 polarization. RGMb is expressed on the surface of naive mouse T cells, macrophages, neutrophils and dendritic cells. Finally, K113S/RGMb costimulation suppresses Th2-mediated asthma and ameliorates small airway inflammation and lung pathology in an experimental mouse model. Our findings indicate that RGMb is a costimulatory receptor for B7-DC. These findings from the K113S variant provide not only a possible explanation for the B7-DC-triggered contradictory effects on T-cell responses, but also a novel approach to investigate the B7-DC/PD-1/RGMb axis. Recombinant K113S or its derivatives could potentially be developed as an agonist for RGMb to costimulate the Th1 response without triggering PD-1-mediated T-cell inhibition.

Protective effect of ATP on skeletal muscle satellite cells damaged by H2O2.

This study investigated the protective effect of ATP on skeletal muscle satellite cells damaged by H2O2in neonatal rats and the possible mechanism. The skeletal muscle satellite cells were randomly divided into four groups: normal group, model group (cells treated with 0.1 mmol/L H2O2for 50 s), protection group (cells treated with 16, 8, 4, 2, 1, 0.5, or 0.25 mmol/L ATP for 24 h, and then with 0.1 mmol/L H2O2for 50 s), proliferation group (cells treated with 16, 8, 4, 2, 1, 0.5, or 0.25 mmol/L ATP for 24 h). MTT assay, FITC+PI+DAPI fluorescent staining, Giemsa staining and immunofluorescence were performed to examine cell viability and apoptosis, and apoptosis-related proteins. The results showed that the survival rate of skeletal muscle satellite cells was decreased and the apoptosis rate was increased after H2O2treatment (P<0.01). Different doses of ATP had different effects on skeletal muscle satellite cells damaged by H2O2: the survival rate of muscle satellite cells treated with ATP at 4, 2, or 1 mmol/L was increased. The protective effect was most profound on cells treated with 2 mmol/L ATP. Immunofluorescence showed that ATP could increase the number of Bcl-2-positive cells (P<0.01) and decrease the number of the Bax-positive cells (P<0.01). It was concluded that ATP could protect skeletal muscle satellite cells against H2O2damage in neonatal rats, which may be attributed to the up-regulation of the expression of Bcl-2 and down-regulation of Bax, resulting in the suppression of apoptosis.