Phenformin activates ER stress to promote autophagic cell death via NIBAN1 and DDIT4 in oral squamous cell carcinoma independent of AMPK.

The efficient clinical treatment of oral squamous cell carcinoma (OSCC) is still a challenge that demands the development of effective new drugs. Phenformin has been shown to produce more potent anti-tumor activities than metformin on different tumors, however, not much is known about the influence of phenformin on OSCC cells. We found that phenformin suppresses OSCC cell proliferation, and promotes OSCC cell autophagy and apoptosis to significantly inhibit OSCC cell growth both in vivo and in vitro. RNA-seq analysis revealed that autophagy pathways were the main targets of phenformin and identified two new targets DDIT4 (DNA damage inducible transcript 4) and NIBAN1 (niban apoptosis regulator 1). We found that phenformin significantly induces the expression of both DDIT4 and NIBAN1 to promote OSCC autophagy. Further, the enhanced expression of DDIT4 and NIBAN1 elicited by phenformin was not blocked by the knockdown of AMPK but was suppressed by the knockdown of transcription factor ATF4 (activation transcription factor 4), which was induced by phenformin treatment in OSCC cells. Mechanistically, these results revealed that phenformin triggers endoplasmic reticulum (ER) stress to activate PERK (protein kinase R-like ER kinase), which phosphorylates the transitional initial factor eIF2, and the increased phosphorylation of eIF2 leads to the increased translation of ATF4. In summary, we discovered that phenformin induces its new targets DDIT4 and especially NIBAN1 to promote autophagic and apoptotic cell death to suppress OSCC cell growth. Our study supports the potential clinical utility of phenformin for OSCC treatment in the future.

Cutaneous inflammasome driving ASC / gasdermin-D activation and IL-1ß-secreting macrophages in severe atopic dermatitis.

Atopic dermatitis (AD) is an inflammatory skin disease with intense pruritus, and chronic skin colonization by Staphylococcus aureus. To understand the inflammatory status in AD, we investigated the inflammasome complex, that activates ASC (Apoptosis-associated speck-like protein containing a CARD), caspase-1 and GSDMD (gasdermin-D), and production of IL-1ß and IL-18. We aimed to evaluate the expression of the inflammasome pathway in the skin of adults with AD. Thirty patients with moderate to severe AD and 20 healthy controls were enrolled in the study. We performed the analysis of the inflammasome components NLRP1, NLRP3, AIM-2, IL-1ß, IL-18, Caspase-1, ASC, GSDMD, and CD68 expression (macrophage marker) by immunohistochemistry and immunofluorescence. The main findings included increased expression of NLRP3, NLRP1 and AIM-2 at dermal level of severe AD; augmented IL-18 and IL-1ß expression at epidermis of moderate and severe patients, and in the dermis of severe AD; augmented expression of ASC, caspase-1 and GSDMD in both epidermis and dermis of moderate and severe AD. We detected positive correlation between caspase-1, GSDMD and IL-1ß (epidermis) and caspase-1 (dermis) and AD severity; NLRP3, AIM-2 and IL-1ß, and NLRP3 with IL-18 in the epidermis; ASC, GSDMD and IL-1ß, and NLRP3, AIM-2, caspase-1, and IL-18 in the dermis. We also evidenced the presence of CD68+ macrophages secreting GSDMD, ASC and IL-1ß in moderate and severe AD. Cutaneous macrophages, early detected in moderate AD, have its role in the disease inflammatory mechanisms. Our study indicates a canonical activation pathway of inflammasomes, reinforced by the chronic status of inflammation in AD. The analysis of the inflammasome complex evidenced an imbalance in its regulation, with increased expression of the evaluated components, which is remarkably in severe AD, emphasizing its relevance as potential disease biomarkers and targets for immunomodulatory interventions.

The Neglected Sibling: NLRP2 Inflammasome in the Nervous System.

While classical NOD-like receptor pyrin domain containing protein 1 (NLRP1) and NLRP3 inflammasomal proteins have been extensively investigated, the contribution of NLRP2 is still ill-defined in the nervous system. Given the putative significance of NLRP2 in orchestrating neuroinflammation, further inquiry is needed to gain a better understanding of its connectome, hence its specific targeting may hold a promising therapeutic implication. Therefore, bioinformatical approach for extracting information, specifically in the context of neuropathologies, is also undoubtedly preferred. To the best of our knowledge, there is no review study selectively targeting only NLRP2. Increasing, but still fragmentary evidence should encourage researchers to thoroughly investigate this inflammasome in various animal- and human models. Taken together, herein we aimed to review the current literature focusing on the role of NLRP2 inflammasome in the nervous system and more importantly, we provide an algorithm-based protein network of human NLRP2 for elucidating potentially valuable molecular partnerships that can be the beginning of a new discourse and future therapeutic considerations.

[Chronic intermittent hypoxia activates NLRP1 inflammasome and causes liver injury].

Objective To investigate the liver injury induced by chronic intermittent hypoxia (CIH) activation of NOD-like receptor pyrin domain containing protein 1 (NLRP1) inflammasome. Methods C57BL/6 male mice were randomly divided into control group and CIH group. Mice in CIH group were put into CIH chamber for molding (8 hours a day for 4 weeks). After 4 weeks of molding, liver tissue cells was observed by HE staining, and the levels of alanine aminotransferase (ALT) and aspartate aminotransferase (AST) in serum of mice were detected by kit. The levels of reactive oxygen species (ROS) in liver tissue were detected by dihydroethidine (DHE). The expression and localization of NLRP1, apoptosis speck-like protein containing a caspase activation and recruiting domain (ASC) and caspase-1 were detected by immunohistochemical staining. The protein expressions of NLRP1, ASC, caspase-1, interleukin 1ß (IL-1ß) and tumor necrosis factor α (TNF-α) were detected by Western blot analysis. The serum levels of IL-1ß and TNF-α were detected by ELISA. Results Compared with the control group, the CIH group exhibited significant pathological changes in hepatocytes. Hepatocytes showed signs of rupture and necrosis, accompanied by inflammatory cell aggregation. Furthermore, the levels of ALT, AST, ROS, IL-1ß and TNF-α were elevated, along with increased protein expressions of NLRP1, ASC, caspase-1, IL-1ß and TNF-α. Conclusion CIH causes liver injury by activating NLRP1 inflammasome.

Dab2 (Disabled-2), an adaptor protein, regulates self-renewal of hair follicle stem cells.

Disabled 2 (Dab2), an adaptor protein, is up regulated in the hair follicle stem cells (HFSCs); however, its role in any tissue stem cells has not been studied. In the present study, we have reported that Dab2 conditional knockout (Dab2-cKO) mice exhibited a delay in the HF cycle due to perturbed activation of HFSCs. Further, Dab2-cKO mice showed a reduction in the number of HFSCs and reduced colony forming ability of HFSCs. Dab2-cKO mice showed extended quiescence of HFSCs concomitant with an increased expression of Nfatc1. Dab2-cKO mice showed a decreased expression of anti-aging genes such as Col17a1, decorin, Sirt2 and Sirt7. Dab2-cKO mice did not show full hair coat recovery in aged mice thereby suggesting an accelerated aging process. Overall, we unveil for the first time, the role of Dab2 that regulate activation and self-renewal of HFSCs.

18ß-Glycyrrhetinic acid protects against deoxynivalenol-induced liver injury via modulating ferritinophagy and mitochondrial quality control.

Deoxynivalenol (DON), a widespread mycotoxin, represents a substantial public health hazard due to its propensity to contaminate agricultural produce, leading to both acute and chronic health issues in humans and animals upon consumption. The role of ferroptosis in DON-induced hepatic damage remains largely unexplored. This study investigates the impact of 18ß-glycyrrhetinic acid (GA), a prominent constituent of glycyrrhiza, on DON hepatotoxicity and elucidates the underlying mechanisms. Our results indicate that GA effectively attenuates liver injury inflicted by DON. This was achieved by inhibiting nuclear receptor coactivator 4 (NCOA4)-mediated ferritinophagy and ferroptosis, as well as by adjusting mitochondrial quality control (MQC). Specifically, GA curtails ferritinophagy by diminishing NCOA4 expression without affecting the autophagic flux. At a molecular level, GA binds to and stabilizes programmed cell death protein 4 (PDCD4), thereby inhibiting its ubiquitination and subsequent degradation. This stabilization of PDCD4 leads to the downregulation of NCOA4 via the JNK-Jun-NCOA4 axis. Knockdown of PDCD4 weakened GA's protective action against DON exposure. Furthermore, GA improved mitochondrial function and limited excessive mitophagy and mitochondrial division induced by DON. Disrupting GA's modulation of MQC nullified its anti-ferroptosis effects. Overall, GA offers protection against DON-induced ferroptosis by blocking ferritinophagy and managing MQC. ENVIRONMENTAL IMPLICATION: Food contamination from mycotoxins, is a problem for agricultural and food industries worldwide. Deoxynivalenol (DON), the most common mycotoxins in cereal commodities. A survey in 2023 showed that the positivity rate for DON contamination in food reached more than 70% globally. DON can damage the health of humans whether exposed to high doses for short periods of time or low doses for long periods of time. We have discovered 18ß-Glycyrrhetinic acid (GA), a prominent constituent of glycyrrhiza. Liver damage caused by low-dose DON can be successfully treated with GA. This study will support the means of DON control, including antidotes.

PDCD4 restricts PRRSV replication in an eIF4A-dependent manner and is antagonized by the viral nonstructural protein 9.

As obligate parasites, viruses have evolved multiple strategies to evade the host immune defense. Manipulation of the host proteasome system to degrade specific detrimental factors is a common viral countermeasure. To identify host proteins targeted for proteasomal degradation by porcine reproductive and respiratory syndrome virus (PRRSV), we conducted a quantitative proteomics screen of PRRSV-infected Marc-145 cells under the treatment with proteasome inhibitor MG132. The data revealed that the expression levels of programmed cell death 4 (PDCD4) were strongly downregulated by PRRSV and significantly rescued by MG132. Further investigation confirmed that PRRSV infection induced the translocation of PDCD4 from the nucleus to the cytoplasm, and the viral nonstructural protein 9 (Nsp9) promoted PDCD4 proteasomal degradation in the cytoplasm by activating the Akt-mTOR-S6K1 pathway. The C-terminal domain of Nsp9 was responsible for PDCD4 degradation. As for the role of PDCD4 during PRRSV infection, we demonstrated that PDCD4 knockdown favored viral replication, while its overexpression significantly attenuated replication, suggesting that PDCD4 acts as a restriction factor for PRRSV. Mechanistically, we discovered eukaryotic translation initiation factor 4A (eIF4A) was required for PRRSV. PDCD4 interacted with eIF4A through four sites (E249, D253, D414, and D418) within its two MA3 domains, disrupting eIF4A-mediated translation initiation in the 5'-untranslated region of PRRSV, thereby inhibiting PRRSV infection. Together, our study reveals the antiviral function of PDCD4 and the viral strategy to antagonize PDCD4. These results will contribute to our understanding of the immune evasion strategies employed by PRRSV and offer valuable insights for developing new antiviral targets.IMPORTANCEPorcine reproductive and respiratory syndrome virus (PRRSV) infection results in major economic losses in the global swine industry and is difficult to control effectively. Here, using a quantitative proteomics screen, we identified programmed cell death 4 (PDCD4) as a host protein targeted for proteasomal degradation by PRRSV. We demonstrated that PDCD4 restricts PRRSV replication by interacting with eukaryotic translation initiation factor 4A, which is required for translation initiation in the viral 5'-untranslated region. Additionally, four sites within two MA3 domains of PDCD4 are identified to be responsible for its antiviral function. Conversely, PRRSV nonstructural protein 9 promotes PDCD4 proteasomal degradation in the cytoplasm by activating the Akt-mTOR-S6K1 pathway, thus weakening the anti-PRRSV function. Our work unveils PDCD4 as a previously unrecognized host restriction factor for PRRSV and reveals that PRRSV develops countermeasures to overcome PDCD4. This will provide new insights into virus-host interactions and the development of new antiviral targets.

BAG3 promotes proliferation and migration of arterial smooth muscle cells by regulating STAT3 phosphorylation in diabetic vascular remodeling.

BACKGROUND: Diabetic vascular remodeling is the most important pathological basis of diabetic cardiovascular complications. The accumulation of advanced glycation end products (AGEs) caused by elevated blood glucose promotes the proliferation and migration of vascular smooth muscle cells (VSMCs), leading to arterial wall thickening and ultimately vascular remodeling. Therefore, the excessive proliferation and migration of VSMCs is considered as an important therapeutic target for vascular remodeling in diabetes mellitus. However, due to the lack of breakthrough in experiments, there is currently no effective treatment for the excessive proliferation and migration of VSMCs in diabetic patients. Bcl-2-associated athanogene 3 (BAG3) protein is a multifunctional protein highly expressed in skeletal muscle and myocardium. Previous research has confirmed that BAG3 can not only regulate cell survival and apoptosis, but also affect cell proliferation and migration. Since the excessive proliferation and migration of VSMCs is an important pathogenesis of vascular remodeling in diabetes, the role of BAG3 in the excessive proliferation and migration of VSMCs and its molecular mechanism deserve further investigation. METHODS: In this study, BAG3 gene was manipulated in smooth muscle to acquire SM22αCre; BAG3FL/FL mice and streptozotocin (STZ) was used to simulate diabetes. Expression of proteins and aortic thickness of mice were detected by immunofluorescence, ultrasound and hematoxylin-eosin (HE) staining. Using human aorta smooth muscle cell line (HASMC), cell viability was measured by CCK-8 and proliferation was measured by colony formation experiment. Migration was detected by transwell, scratch experiments and Phalloidin staining. Western Blot was used to detect protein expression and Co-Immunoprecipitation (Co-IP) was used to detect protein interaction. RESULTS: In diabetic vascular remodeling, AGEs could promote the interaction between BAG3 and signal transducer and activator of transcription 3 (STAT3), leading to the enhanced interaction between STAT3 and Janus kinase 2 (JAK2) and reduced interaction between STAT3 and extracellular signal-regulated kinase 1/2 (ERK1/2), resulting in accumulated p-STAT3(705) and reduced p-STAT3(727). Subsequently, the expression of matrix metallopeptidase 2 (MMP2) is upregulated, thus promoting the migration of VSMCs. CONCLUSIONS: BAG3 upregulates the expression of MMP2 by increasing p-STAT3(705) and decreasing p-STAT3(727) levels, thereby promoting vascular remodeling in diabetes. This provides a new orientation for the prevention and treatment of diabetic vascular remodeling.

[Huangpu Tongqiao Capsule improves cognitive impairment in rats with Wilson disease by inhibiting endoplasmic reticulum stress-mediated apoptosis pathway].

OBJECTIVE: To investigate the neuroprotective effect of Huangpu Tongqiao Capsule (HPTQ) in a rat model of Wilson disease (WD) and explore the underlying mechanisms. METHODS: SD rat models of WD were established by feeding of coppersupplemented chow diet and drinking water for 12 weeks, and starting from the 9th week, the rats were treated with low-, moderate- and high-dose HPTQ, penicillamine, or normal saline by gavage on a daily basis for 3 weeks. Copper levels in the liver and 24-h urine of the rats were detected, and their learning and memory abilities were evaluated using Morris water maze test. HE staining was used to observe morphological changes of CA1 region neurons in the hippocampus, and neuronal apoptosis was detected with TUNEL staining. Hippocampal expressions of endoplasmic reticulum stress (ERS)-mediated apoptosis pathway-related proteins GRP78, CHOP, caspase-12, cleaved caspase-9, and cleaved caspase-3 at both the mRNA and protein levels were detected using RT-qPCR, immunofluorescence assay or Western blotting. RESULTS: Compared with normal control rats, the rat models with copper overload-induced WD exhibited significantly increased copper levels in both the liver and 24-h urine, impaired learning and memory abilities, obvious hippocampal neuronal damage in the CA1 region and increased TUNEL-positive neurons (P<0.01), with also lowered mRNA and protein expressions of GRP78, CHOP, caspase-12, cleaved caspase-9, and cleaved caspase-3 in the hippocampus (all P<0.01). Treatments with HPTQ and penicillamine significantly lowered copper level in the liver but increased urinary copper level, improved learning and memory ability, alleviated neuronal damage and apoptosis in the hippocampus, and decreased hippocampal expressions of GRP78, CHOP, caspase-12, cleaved caspase-9, and cleaved caspase-3 in the rat models (P<0.01 or 0.05). CONCLUSION: HPTQ Capsule has neuroprotective effects in rat models of WD possibly by inhibiting ERS-mediated apoptosis pathway.

Progress on the relationship between tumor suppressor PDCD4 and diseases based on the analysis of structural characteristics.

The tumor suppressor programmed cell death 4 (PDCD4) is downregulated in various tumor tissues indicating poor prognosis. PDCD4 is the first protein found to resist tumor transformation, invasion, and metastasis by inhibiting translation. The functions of PDCD4 dependent on its structures are affected by extracellular signals. It regulates tumor-related proteins through a variety of mechanisms, especially involved in two major signaling pathways, PI3K-Akt-mTOR and MAPK. By analyzing the relationship between the structures, functions and diseases of PDCD4, this review summarizes the roles of PDCD4 in several physiological processes and diseases such as apoptosis, autophagy, tumor, and inflammation in recent years, thereby providing insights for the study of the signaling pathways of PDCD4 and related proteins and the treatment of diseases targeting them.

Kinetochore scaffold 1 downregulation suppressed the development of non-small cell lung cancer by inactivating the phosphatidylinositol 3 kinase/protein kinase B (AKT)/nuclear factor-kappa B pathway.

Kinetochore scaffold 1 (KNL1) is indispensable for generating motile micro-tubule attachments and isolating chromosomes. KNL1 is highly expressed in multiple middle-route tissues and promotes tumor development. However, how it functions in non-small cell lung cancer (NSCLC) is unclear. Real-time quantitative PCR (RT-qPCR) and Western blotting (WB) were used to determine KNL1 expression in NSCLC tissues and cells. The sh-KNL1 or oe-KNL1 was transfected into NSCLC cells. The colony formation assay, cell counting kit-8 (CCK-8) assay, and flow cytometry were used to evaluate cell proliferation and apoptosis. A transwell assay was used to monitor invasion and migration. The CCK-8 assay was used to measure NSCLC cell sensitivity to chemotherapy drugs. WB confirmed the protein levels of apoptosis-related proteins, cell cycle-associated proteins, and the phosphatidylinositol 3 kinase (PI3K)/protein kinase B (AKT)/nuclear factor kappaB (NF-κB) pathway. A PI3K/AKT/NF-κB pathway inhibitor was used to intervene in NSCLC cell transfection along with oe-KNL1, thus revealing the function of the pathway in carcinogenicity mediated by KNL1. In result KNL1 expression was substantially increased in NSCLC tissues and cells. High-level KNL1 expression is related to the poor prognosis of NSCLC patients. KNL1 silencing bolstered promoted NSCLC cell apoptosis and inhibited proliferation, cell cycle progression, invasion, and EMT, whereas KNL1 silencing had the opposite effect. KNL1 knockdown increased NSCLC cell sensitivity to chemical drugs. KNL1 promoted PI3K/AKT/NF-κB pathway activation, while PI3K/AKT/NF-κB pathway inhibition weakened the procancer effect mediated by KNL1 overexpression but had little influence on KNL1 levels. We conclude that KNL1 activates the PI3K/AKT/NF-κB pathway to increase NSCLC progression and attenuate NSCLC sensitivity to chemotherapy drugs.

STAT1 mediated downregulation of the tumor suppressor gene PDCD4, is driven by the atypical cadherin FAT1, in glioblastoma.

STAT1 (Signal Transducer and Activator of Transcription 1), belongs to the STAT protein family, essential for cytokine signaling. It has been reported to have either context dependent oncogenic or tumor suppressor roles in different tumors. Earlier, we demonstrated that Glioblastoma multiforme (GBMs) overexpressing FAT1, an atypical cadherin, had poorer outcomes. Overexpressed FAT1 promotes pro-tumorigenic inflammation, migration/invasion by downregulating tumor suppressor gene, PDCD4. Here, we demonstrate that STAT1 is a novel mediator downstream to FAT1, in downregulating PDCD4 in GBMs. In-silico analysis of GBM databases as well as q-PCR analysis in resected GBM tumors showed positive correlation between STAT1 and FAT1 mRNA levels. Kaplan-Meier analysis showed poorer survival of GBM patients having high FAT1 and STAT1 expression. SiRNA-mediated knockdown of FAT1 decreased STAT1 and increased PDCD4 expression in glioblastoma cells (LN229 and U87MG). Knockdown of STAT1 alone resulted in increased PDCD4 expression. In silico analysis of the PDCD4 promoter revealed four putative STAT1 binding sites (Site1-Site4). ChIP assay confirmed the binding of STAT1 to site1. ChIP-PCR revealed decrease in the binding of STAT1 on the PDCD4 promoter after FAT1 knockdown. Site directed mutagenesis of Site1 resulted in increased PDCD4 luciferase activity, substantiating STAT1 mediated PDCD4 inhibition. EMSA confirmed STAT1 binding to the Site 1 sequence. STAT1 knockdown led to decreased expression of pro-inflammatory cytokines and EMT markers, and reduced migration/invasion of GBM cells. This study therefore identifies STAT1 as a novel downstream mediator of FAT1, promoting pro-tumorigenic activity in GBM, by suppressing PDCD4 expression.

Study on the role of CCM3 gene and lead exposure induced neurotoxicity through neurovascular units.

This study aimed to determine the toxic effects of vascular CCM3 gene deficiency and lead (Pb) exposure on the nervous system. Lentiviral transfection was performed to generate a stable strain of brain microvascular endothelial cells with low CCM3 expression. MTT assay assessed the survival rate of cells exposed to Pb, determining the dose and duration of Pb exposure in vitro. Proteomic analysis was performed on the differentially expressed proteins in bEnd3 and HT22 cells and flow cytometry was used to detect cell apoptosis. Finally, urine samples from pregnant and postpartum women were subjected to ICP-MS to detect Pb levels and HPLC to detect neurotransmitter metabolites. Based on the proteomic analysis of bEnd3 (CCM3-/-) cells co-cultured with HT22 cells, it was determined that HT22 cells and CCM3 genes interfered with bEnd3 cell differential proteins,2 including apoptosis and ferroptosis pathways. Electron microscopy observation, ICP-MS iron ion loading detection, and WB determination of protein GPX4 expression confirmed that HT22 cells undergo apoptosis, while bEnd3 cells undergo multiple pathways of iron death and apoptosis regulation. Furthermore, a linear regression model showed the interaction between maternal urine Pb levels, the rs9818496 site of the CCM3 SNP in peripheral blood DNA, and the concentration of the neurotransmitter metabolite 5-HIAA in maternal urine (F=4.198, P < 0.05). bEnd3 cells with CCM3 gene deficiency can induce HT22 cell apoptosis through iron death and apoptosis pathways under Pb exposure in a combined cell culture Pb exposure model, and CCM3 gene deficiency in endothelial cells and Pb exposure interacts with neural cell HT22. Epidemiological studies on maternal and newborn infants further confirmed the interaction between urine Pb levels in mothers and the SNP rs9818496 site of the CCM3 gene in peripheral blood DNA.

Direct Salmonella injection into enteroid cells allows the study of host-pathogen interactions in the cytosol with high spatiotemporal resolution.

Intestinal epithelial cells (IECs) play pivotal roles in nutrient uptake and in the protection against gut microorganisms. However, certain enteric pathogens, such as Salmonella enterica serovar Typhimurium (S. Tm), can invade IECs by employing flagella and type III secretion systems (T3SSs) with cognate effector proteins and exploit IECs as a replicative niche. Detection of flagella or T3SS proteins by IECs results in rapid host cell responses, i.e., the activation of inflammasomes. Here, we introduce a single-cell manipulation technology based on fluidic force microscopy (FluidFM) that enables direct bacteria delivery into the cytosol of single IECs within a murine enteroid monolayer. This approach allows to specifically study pathogen-host cell interactions in the cytosol uncoupled from preceding events such as docking, initiation of uptake, or vacuole escape. Consistent with current understanding, we show using a live-cell inflammasome reporter that exposure of the IEC cytosol to S. Tm induces NAIP/NLRC4 inflammasomes via its known ligands flagellin and T3SS rod and needle. Injected S. Tm mutants devoid of these invasion-relevant ligands were able to grow in the cytosol of IECs despite the absence of T3SS functions, suggesting that, in the absence of NAIP/NLRC4 inflammasome activation and the ensuing cell death, no effector-mediated host cell manipulation is required to render the epithelial cytosol growth-permissive for S. Tm. Overall, the experimental system to introduce S. Tm into single enteroid cells enables investigations into the molecular basis governing host-pathogen interactions in the cytosol with high spatiotemporal resolution.

PDCD4-induced oxidative stress through FGR/NF-κB axis in rectal cancer radiotherapy-induced AKI.

This study aimed to investigate the molecular mechanism of the effect of PDCD4 on radiotherapy-induced acute kidney injury (AKI) in rectal cancer through the regulation of FGR/NF-κB signaling. Differentially expressed genes were identified using Gene Expression Omnibus (GEO) datasets (GSE90627 for rectal cancer and GSE145085 for AKI) and R software. The human renal tubular epithelial cell line, HK-2, was used to establish an in vitro model of radiotherapy-induced AKI. RT-qPCR and western blotting were used to detect gene and protein expression levels, respectively. Cell proliferation and apoptosis were assessed using the CCK-8 assay and flow cytometry, respectively. The malondialdehyde and superoxide dismutase levels in the cell culture supernatants were determined. Additionally, an in vivo AKI model was established using BALB/c mice, and kidney tissue morphology, expression of the renal injury molecule KIM-1, apoptosis of renal tubular cells, and TAS and TOS in serum were evaluated. Bioinformatics analysis revealed the upregulated expression of PDCD4 in AKI. In vitro experiments demonstrated that PDCD4 induced apoptosis in renal tubular cells by promoting FGR expression, which activated the NF-κB signaling pathway and triggered an oxidative stress response. In vivo animal experiments confirmed that PDCD4 promoted oxidative stress response and radiotherapy-induced AKI through the activation of the FGR/NF-κB signaling pathway. Silencing PDCD4 attenuated radiotherapy-induced AKI. Our findings suggest that PDCD4 may induce radiotherapy-induced AKI in rectal cancer by promoting FGR expression, activating the NF-κB signaling pathway, and triggering an oxidative stress response.

Nlrp2 deletion ameliorates kidney damage in a mouse model of cystinosis.

Cystinosis is a rare autosomal recessive disorder caused by mutations in the CTNS gene that encodes cystinosin, a ubiquitous lysosomal cystine/H+ antiporter. The hallmark of the disease is progressive accumulation of cystine and cystine crystals in virtually all tissues. At the kidney level, human cystinosis is characterized by the development of renal Fanconi syndrome and progressive glomerular and interstitial damage leading to end-stage kidney disease in the second or third decade of life. The exact molecular mechanisms involved in the pathogenesis of renal disease in cystinosis are incompletely elucidated. We have previously shown upregulation of NLRP2 in human cystinotic proximal tubular epithelial cells and its role in promoting inflammatory and profibrotic responses. Herein, we have investigated the role of NLRP2 in vivo using a mouse model of cystinosis in which we have confirmed upregulation of Nlrp2 in the renal parenchyma. Our studies show that double knock out Ctns-/- Nlrp2-/- animals exhibit delayed development of Fanconi syndrome and kidney tissue damage. Specifically, we observed at 4-6 months of age that animals had less glucosuria and calciuria and markedly preserved renal tissue, as assessed by significantly lower levels of inflammatory cell infiltration, tubular atrophy, and interstitial fibrosis. Also, the mRNA expression of some inflammatory mediators (Cxcl1 and Saa1) and the rate of apoptosis were significantly decreased in 4-6-month old kidneys harvested from Ctns-/- Nlrp2-/- mice compared to those obtained from Ctns-/-mice. At 12-14 months of age, renal histological was markedly altered in both genetic models, although double KO animals had lower degree of polyuria and low molecular weight proteinuria and decreased mRNA expression levels of Il6 and Mcp1. Altogether, these data indicate that Nlrp2 is a potential pharmacological target for delaying progression of kidney disease in cystinosis.

BAG3 localizes to mitochondria in cardiac fibroblasts and regulates mitophagy.

The co-chaperone Bcl2-associated athanogene 3 (BAG3) is a central node in protein quality control in the heart. In humans and animal models, decreased BAG3 expression is associated with cardiac dysfunction and dilated cardiomyopathy. Although previous studies focused on BAG3 in cardiomyocytes, cardiac fibroblasts are also critical drivers of pathologic remodeling. Yet, the role of BAG3 in cardiac fibroblasts is almost completely unexplored. Here, we show that BAG3 is expressed in primary rat neonatal cardiac fibroblasts and preferentially localizes to mitochondria. Knockdown of BAG3 reduces mitophagy and enhances fibroblast activation, which is associated with fibrotic remodeling. Heat shock protein 70 (Hsp70) is a critical binding partner for BAG3 and inhibiting this interaction in fibroblasts using the drug JG-98 decreased autophagy, decreased mitofusin-2 expression, and disrupted mitochondrial morphology. Together, these data indicate that BAG3 is expressed in cardiac fibroblasts, where it facilitates mitophagy and promotes fibroblast quiescence. This suggests that depressed BAG3 levels in heart failure may exacerbate fibrotic pathology, thus contributing to myocardial dysfunction through sarcomere-independent pathways.NEW & NOTEWORTHY We report BAG3's localization to mitochondria and its role in mitophagy for the first time in primary ventricular cardiac fibroblasts. We have also collected the first evidence showing that loss of BAG3 increases cardiac fibroblast activation into myofibroblasts, which are major drivers of cardiac fibrosis and pathological remodeling during heart disease.

A novel C-type lectin protein (BjCTL5) interacts with apoptosis stimulating proteins of p53 (ASPP) to activate NF-κB signaling pathway in primitive chordate.

C-type lectin proteins (CTLs), a group of pattern recognition receptors (PRRs), play pivotal roles in immune responses. However, the signal transduction and regulation of CTLs in cephalochordates have yet to be explored. In this study, we examined the composition of CTLs in Branchiostoma japonicum, identifying a total of 272 CTLs. These CTLs underwent further analysis concerning domain arrangement, tandem and segmental duplication events. A multidomain C-type lectin gene, designated as BjCTL5, encompassing CLECT, KR, CUB, MAM, and SR domains, was the focal point of our investigation. BjCTL5 exhibits ubiquitous expression across all detected tissues and is responsive to stimulation by LPS, mannose, and poly (I:C). The recombinant protein of BjCTL5 can bind to Escherichia coli and Staphylococcus aureus, inducing their agglutination and inhibiting the proliferation of S. aureus. Yeast two-hybrid, CoIP, and confocal immunofluorescence experiments revealed the interaction between BjCTL5 and apoptosis-stimulating proteins of p53, BjASPP. Intriguingly, BjCTL5 was observed to induce the luciferase activity of the NF-κB promoter in HEK293T cells. These results suggested a potential interaction between BjCTL5 and BjASPP, implicating that they involve in the activation of the NF-κB signaling pathway, which provides an evolutionary viewpoint on NF-κB signaling pathway in primitive chordate.

Multifaceted roles of PDCD6 both within and outside the cell.

Programmed cell death protein 6 (PDCD6) is an evolutionarily conserved Ca2+-binding protein. PDCD6 is involved in regulating multifaceted and pleiotropic cellular processes in different cellular compartments. For instance, nuclear PDCD6 regulates apoptosis and alternative splicing. PDCD6 is required for coat protein complex II-dependent endoplasmic reticulum-to-Golgi apparatus vesicular transport in the cytoplasm. Recent advances suggest that cytoplasmic PDCD6 is involved in the regulation of cytoskeletal dynamics and innate immune responses. Additionally, membranous PDCD6 participates in membrane repair through endosomal sorting complex required for transport complex-dependent membrane budding. Interestingly, extracellular vesicles are rich in PDCD6. Moreover, abnormal expression of PDCD6 is closely associated with many diseases, especially cancer. PDCD6 is therefore a multifaceted but pivotal protein in vivo. To gain a more comprehensive understanding of PDCD6 functions and to focus and stimulate PDCD6 research, this review summarizes key developments in its role in different subcellular compartments, processes, and pathologies.