Low Expression Research Articles

Sex-specific mechanisms of cerebral microvascular BKCa dysfunction in a mouse model of Alzheimer's disease.

Cerebrovascular dysfunction occurs in Alzheimer's disease (AD), impairing hemodynamic regulation. Large conductance Ca2+-activated K+ channels (BKCa) regulate cerebrovascular reactivity and are impaired in AD. BKCa activity depends on intracellular Ca2+ (Ca2+ sparks) and nitro-oxidative post-translational modifications. However, whether these mechanisms underlie BKCa impairment in AD remains unknown. Cerebral arteries from 5x-FAD and wild-type (WT) littermates were used for molecular biology, electrophysiology, ex vivo, and in vivo experiments. Arterial BKCa activity is reduced in 5x-FAD via sex-dependent mechanisms: in males, there is lower BKα subunit expression and less Ca2+ sparks. In females, we observed reversible nitro-oxidative modification of BKCa. Further, BKCa is involved in hemodynamic regulation in WT mice, and its dysfunction is associated with vascular deficits in 5x-FAD. Our data highlight the central role played by BKCa in cerebral hemodynamic regulation and that molecular mechanisms of its impairment diverge based on sex in 5x-FAD. Cerebral microvascular BKCa dysfunction occurs in both female and male 5x-FAD. Reduction in BKα subunit protein and Ca2+ sparks drive the dysfunction in males. Nitro-oxidative stress is present in females, but not males, 5x-FAD. Reversible nitro-oxidation of BKα underlies BKCa dysfunction in female 5x-FAD.

RAB37 suppresses the EMT, migration and invasion of gastric cancer cells by mediating autophagic degradation of β-catenin.

Gastric cancer, characterized by its high morbidity and mortality rates, exhibits low levels of RAB37. The role and molecular mechanisms of RAB37, a small GTPase, in the pathogenesis of gastric cancer are still unclear. We assessed RAB37 expression in gastric cancer cells using quantitative Polymerase Chain Reaction (qPCR), Western blot, and immunohistochemical staining (IHC), and analyzed EMT marker proteins and autophagy changes via Western blot, immunofluorescence (IF), and transmission electron microscopy (TEM). Co-immunoprecipitation (co-IP) was used to identify protein-protein interactions. We studied the migration and invasion of gastric cancer cells using wound healing and transwell assays in vitro and a mouse pulmonary metastasis model in vivo. Overexpression of RAB37 suppressed EMT, invasion, and migration while enhancing autophagy in gastric cancer cells, which was dependent on its GTPase activity. However, all these effects could be reversed by the autophagy inhibitor chloroquine. Regarding the molecular mechanism, RAB37 strengthened the interaction between p62 and β-catenin, which consequently enhanced the p62-mediated autophagic degradation of β-catenin. Furthermore, RAB37 curbed the pulmonary metastasis of both general and cisplatin-resistant gastric cancer cells. The low level of RAB37 reduces interaction between p62 and β-catenin and then the autophagic degradation of β-catenin, thereby promoting the EMT, invasion, and migration in gastric cancer cells. The low expression of RAB37 in gastric cancer suggests a potential therapeutic target, especially for cisplatin-resistant gastric cancer.

Unveiling the shared genes between systemic sclerosis and lung cancer

The risk of lung cancer is significantly increased in patients with systemic sclerosis (SSc), yet the specific genes underlying this association remain unexplored. Our study aims to identify genes shared by SSc and lung cancer. We identified differentially expressed genes (DEGs) from SSc and lung adenocarcinoma (LUAD) datasets (SSc: GSE95065, LUAD: GSE136043) in the GEO database. We found shared genes by intersecting top genes in protein–protein interaction networks by the STRING database. The area under the ROC curve (AUC) was calculated for each shared gene in validation datasets (SSc: GSE231692; LUAD: GSE43458), identifying PRKG2 as the core shared gene. We used the UALCAN platform to assess PRKG2 expression in LUAD patients at various stages and lymph node metastasis states, and compared disease-free survival (DFS) between low and high PRKG2 expression LUAD groups. PRKG2 was overexpressed in A549 cells to study its impact on lung cancer cell proliferation and invasion in vitro. We identified seven shared genes (SCN7A, AGTR1, WIF1, PRKG2, LTF, AQP4, COL10A1), with the AUC for PRKG2 exceeding 0.93 in both diseases (SSc AUC = 0.973; LUAD AUC = 0.939). The PRKG2 expression levels of LUAD patients with different clinical stages and lymph node metastasis states were consistently lower than those observed in normal individuals. The DFS of LUAD patients in the high PRKG2 expression group was higher than that in the low expression group (p = 0.028). In vitro experiments confirmed elevated PRKG2 expression inhibits the proliferation and invasion of lung cancer cells. PRKG2 is one of the genes shared by SSc and lung cancer, affecting the proliferation and invasion of lung cancer cells.

The Hansen's baccata #2 gene Rvi12_Cd5 confers scab resistance to the susceptible apple cultivar "Gala Galaxy".

To enhance the breeding of new scab-resistant apple cultivars, a comprehensive understanding of the mechanisms governing major scab resistance genes is essential. Rvi12_Cd5 was previously identified as the best candidate gene for the Rvi12 scab resistance of the crab apple "Hansen's baccata #2" by gene prediction and in silico analysis. In the present study, Rvi12_Cd5 was used to transform the scab-susceptible apple cultivar "Gala Galaxy." Two constructs were prepared: the first carrying Rvi12_Cd5 under the control of a 35S promoter and E9 terminator, and the second carrying Rvi12_Cd5 under the control of its native promoter and terminator. All the transgenic lines were analyzed for T-DNA integration, copy number, and expression of Rvi12_Cd5 and phenotypically evaluated for scab resistance. The "Gala Galaxy" lines carrying the 35S promoter expressed Rvi12_Cd5 at a high level, showing partial to high resistance against a mixed inoculum of Venturia inaequalis, with symptoms ranging from class 0 to 3b on the Chevalier scale. The transgenic lines carrying the native promoter showed a lower expression of Rvi12_Cd5 compared with the 35S lines. Nevertheless, the low expression was sufficient to induce a resistance level comparable to that of the transgenic lines carrying the 35S promoter. These results indicate that Rvi12_Cd5 confers scab resistance to a susceptible apple cultivar and that even a low level of gene transcript can trigger a plant response to V.inaequalis infection. After HcrVf2 and Vr2-C, Rvi12_Cd5 is the third major apple scab resistance gene being functionally proven.

Stromal immune cells expression of Siglec-15 is associated with lower T stage and better prognosis of urinary bladder cancer

IntroductionSialic acid-binding immunoglobulin-like lectin 15 (Siglec-15) is a novel immune checkpoint, similar to programmed death-ligand (PD-L1), and has emerged as a potential target for cancer immunotherapy. Until recently, little was known about the expression and role of Siglec-15 in bladder cancer (BC).MethodsIn this study, we used immunohistochemical staining to assess the expression of Siglec-15 and PD-L1 in 69 primary BC samples and analyzed their relationship with clinicopathologic characters and prognosis.ResultsThe expression rates of Siglec-15 in the tumor cells, stromal immune cells, and both the tumor and stromal cells were 84.1% (58/69), 50.7% (35/69), and 44.9% (31/69), respectively. The PD-L1 expression rate was 52.2% (36/69), with a positive rate of 17.4% (12/69). PD-L1 expression was inversely correlated with Siglec-15 expression, but the statistical significance was not achieved (P = 0.072). Low stromal Siglec-15 expression was associated with advanced tumor stage (P = 0.010). PD-L1 expression was associated with tumor stage (P = 0.008) and perineural invasion (PNI) (P = 0.048). Kaplan-Meier survival curves showed that stromal Siglec-15 expression was associated with a better prognosis (P = 0.012), although it was not an independent prognostic factor after multivariate analysis (P = 0.236) .DiscussionThis study revealed a high expression rate of Siglec-15 in BC and may provide valuable insights for patient selection in future clinical trials.

Molecular Markers in Embryo Non-Development: Analysis of Gene Expressions (Ki-67, hTERT, HIF-1α) in Spent Embryo Culture Medium

An embryo culture medium is a specialized set of ambient conditions, technological equipment, and nutrients that embryos require to grow properly. We aimed to investigate the Ki-67, hTERT, and HIF-1α gene expression differences between developing and non-developing embryos in spent embryo culture medium. Ki-67, hTERT, and HIF-1α gene expressions were determined from the spent embryo culture medium containing developing and non-developing embryos of 20 normoresponder patients admitted to the Bahçeci Umut IVF Center. An increase in hTERT gene expression (p < 0.05) and a decrease in HIF-1α gene expression (p < 0.001) were observed in mediums of developing compared to the non-developing embryos. No difference was observed in Ki-67 gene expression (p > 0.05). While there was a correlation between Ki-67 and HIF-1α genes in the non-growing group (r < 0.01); no correlation was observed in the developing group (r > 0.05). Both normoresponder groups will be similar in terms of proliferation rate. The low HIF-1α expression that observed high telomerase activity in embryo development maintains continuity and avoids mechanisms that result in cell death. A molecular study of the embryo development in patients with similar characteristics may help to understand the pathogenesis of the disease and establish a diagnosis and specific treatment.

Prognostic, oncogenic roles, and pharmacogenomic features of AMD1 in hepatocellular carcinoma

Background AMD1 is the gene encoding S-adenosylmethionine decarboxylase 1. How AMD1 affects the prognosis of hepatocellular carcinoma (HCC) patients is unclear.MethodsUsing the Cancer Genome Atlas (TCGA) liver hepatocellular carcinoma datasets, gene enrichment and immunological traits were compared between groups with high and low AMD1 expression. After altering AMD1 expression in HCC cells, cell viability, the clonal formation rate, and migration and invasion ability were detected. Univariate Cox regression analysis and Pearson correlation were used to screen for AMD1-related genes (ARGs). Multidimensional bioinformatic algorithms were utilized to establish a risk score model for ARGs.Results AMD1 expression was notably increased in the majority of cancer types. High AMD1 expression was associated with adverse outcomes and poorer immunotherapy response in HCC patients. AMD1 exhibited higher expression levels in HCC cell lines. The efficient inhibition of HCC cell proliferation, migration, and invasion in vitro can be achieved through the downregulation of AMD1. The AMD1-related risk score was constructed with the expression of 9 ARGs, and demonstrated high predictive efficacy in multiple validation cohorts. Patients with high risk scores exhibited greater resistance to classical chemotherapy drugs. The nomogram, which consists of age, stage, and the AMD1-related risk score, was used to calculate the probability of survival for each individual.ConclusionThe present study indicates that AMD1 functions as a potential role in HCC progression and may serve as a therapeutic target in HCC. This study constructed a novel AMD1-related scoring system for predicting the prognosis and treatment responsiveness of patients with HCC, enabling the prediction of prognosis and identification of potential treatment targets.

Significant association of miRNA 34a with BRCA1 expression in pancreatic ductal adenocarcinoma: an insight on miRNA regulatory pathways in the Pakistani population

BackgroundPancreatic Ductal Adenocarcinoma (PDAC) is among the most aggressive cancers, characterized by high mortality rates. Studies on various cancers across the globe indicate that regulatory miRNAs play a vital role in cellular signaling. However, the expression and interactions of these miRNAs in the Pakistani patients with PDAC is yet to be explored. Here, we aim to investigate a panel of four regulatory miRNAs (miRNA 34a, 30b, 142 and 137) in PDAC and their interaction with selected target proteins in the signaling pathway (KRAS, p53, BRCA1, APC).MethodsWe conducted a study on 109 PDAC patients to analyze the selected miRNAs and protein targets. Formalin Fixed Paraffin Embedded (FFPE) tumor samples were obtained from the hospital’s department of histopathology. After confirmation of diagnosis and appropriate tumor content, tissues were processed for RNA extraction. Based on the acceptable quality and quantity of RNA, 43 samples were proceeded for qRT-PCR. Relative expression of the miRNAs was determined through 2−[ΔΔCt] method. Further, FFPE tumor blocks were used to perform tissue sectioning followed by immunohistochemistry experiments. Stained slides were scored independently by two pathologists according to set criteria.ResultsExpression profiles revealed that miRNA 34a, 30b, and 142 showed high expression in approximately 69–70% of cases, while miRNA 137 had a lower high expression frequency (53.4%). Among protein biomarkers, KRAS, BRCA1, and APC were predominantly expressed, with high expression levels observed in 79.1%, 69.8%, and 51.2% of cases, respectively, whereas p53 showed positive expression in only 34.9% of cases. Statistical analysis showed that expression of miRNA 34a was significantly associated with the expression of BRCA1 (p = 0.034). No significant associations were observed for KRAS, p53, or APC with the selected miRNAs. Moreover, the expression of miRNA 34a independently showed significant association with miRNA 30b (p = 0.000) and miRNA 137 (p = 0.001). None of the miRNA showed an association with the overall survival, patient demographics or the clinicopathological characteristics.ConclusionOur study highlights a potential bi-directional regulatory relationship between BRCA1 and miRNA 34a, suggesting that miRNA 34a may both respond to and influence BRCA1 activity within cellular signaling pathways. This complex interaction points to a layered regulatory network that could play a crucial role in tumor suppression in PDAC, underscoring the therapeutic potential of targeting this miRNA-protein crosstalk.

Diagnostic value of multi-tumor-associated autoantibody expression in esophageal squamous cell carcinoma and correlation of clinical features

ObjectiveThis study aimed to investigate the diagnostic value of 7-tumor associated autoantibodies (7-TAAB) and to evaluate the relationship between 7-TAAB and clinical features in esophageal squamous cell carcinoma (ESCC), which can be used to guide clinical diagnosis and treatment and achieve its clinical value.Methods(1) Blood specimens were collected from patients with ESCC who had not previously received antitumor therapy (ESCC group) and those who had normal medical check-ups in the hospital during the same period (control group). The concentrations of 7-TAAB (P53, PGP9.5, SOX2, GAGE7, GBU4-5, MAGE A1, and CAGE) in serum were determined by enzyme-linked immunosorbent assay. The concentrations of 7-TAAB were compared between the ESCC and control groups, and the positive rate of 7-TAAB was calculated to determine the sensitivity, specificity, and accuracy of 7-TAAB. The diagnostic value of 7-TAAB was analyzed using the receiver operating characteristic (ROC) curve. (2) The clinical data of patients with ESCC were collected and the correlation between the rate of 7-TAAB and clinical features was analyzed.Results(1) The serum levels and positivity rates of five antibodies (PGP9.5, SOX2, GBU4-5, MAGE-A1, and CAGE) were higher in the ESCC group than in the control group (P < 0.05) and the positive expression rate of the combined serum 7-TAAB in the ESCC group was significantly higher than that in the control group (P < 0.05). (2) The sensitivity of single antibody detection was 4.20%–17.65%, with a specificity of 96.49%–100%, and accuracy of 51.07%–57.94%. The sensitivity of 7-TAAB combined detection was 49.58%, the specificity was 92.98%, and the accuracy was 70.81%. (3) The ROC curve showed that the 7-TAAB combined test had a certain diagnostic value for ESCC and that its diagnostic efficacy was significantly higher than that of the single autoantibody tests. The diagnostic efficacy of the combined test with the remaining five antibodies (PGP9.5, SOX2, GBU4-5, MAGE-A1, and CAGE) was similar to that of the 7-TAAB combined test after eliminating the two antibodies with low expression rates. (4) Univariate analysis revealed significant differences in the positive expression rates of the 7-TAAB combination test in terms of age, hemoglobin level, albumin level, tumor location, tumor length, lymph node stage, and tumor clinical stage (P < 0.05), and multivariate analysis revealed that age and lymph node stage were independent factors affecting antibody expression.ConclusionThe multi-tumor-associated autoantibody combination test not only has a good auxiliary diagnostic value but also closely correlates with the clinical features of ESCC.

Low Expression of FAM96B is Associated with Poor Prognosis in Hepatocellular Carcinoma.

Recently, studies on FAM96B functions mainly focused on its role in maintaining the normal physiological function of cells. However, the clinical implications of FAM96B in hepatocellular carcinoma (HCC) are still unclear. FAM96B mRNA expression was detected in human HCC tissues and the matched nontumorous tissues by quantitative real-time reverse transcription (qRT-PCR) and then validated in The Cancer Genome Atlas (TCGA) database. Immunohistochemistry assay (IHC) was performed on all 137 cases of HCC samples to examine the protein level of FAM96B. Subsequently, the associations between FAM96B expression and clinicopathological parameters and prognosis were further analyzed. The mRNA level of FAM96B was found to be significantly lower in HCC tissues compared to non-tumorous tissues, as observed in both the local hospital and TCGA database.Immunohistochemistry assay analysis revealed a decrease in FAM96B expression in 78 out of 137 cases, which was significantly associated with larger tumor size, higher Barcelona clinic liver cancer or Child stage, and early distant metastasis. Patients with low FAM96B levels tended to have an unfavorable disease-free and overall survival. Moreover, FAM96B was identified as an independent predictor of patient prognosis in both univariate and multivariate survival analyses. Mechanistically, FAM96B was found to inhibit cancer progression by inducing apoptosis in liver cancer cells and inhibiting their growth. Our findings provide the first evidence suggesting the involvement of FAM96B in the progression of HCC. Additionally, FAM96B could potentially serve as a marker for tumor recurrence and prognosis in HCC patients.

Evaluating toxicity and level of DNA damage in human fetal lung cells upon exposure to 5-methyltetrahydrofolate (bioactive folate).

Background: Folic acid (FA) supplementation is widely regarded as a key nutritional intervention during pregnancy due to its protective effect against neural tube defects. Recent research has reported FA supplementation outcomes on offspring's health, with increased incidences of allergy/respiratory problems. Aim: This study evaluates if increased levels of 5-methyltetrahydrofolate (5-MTHF) are associated with DNA modification, leading to disruption of cell proliferation in fetal lung cells and increasing susceptibility to asthma. Methods: Two fetal lung cells, MRC5 and IMR90, were treated with nine concentrations of 5-MTHF for six time points. Cell viability was evaluated using Trypan Blue staining. Flow cytometry analysis to quantify DNA content in cells was done with a propidium iodide stain. Followed by 1.6 mM glutathione treatment to alleviate the oxidative stress caused by 5-MTHF. A quantitative test for DNA damage was executed using neutral and alkaline comet assay. Gene expression study for five genes namely MTR, MTHFD1, XRCC1, Pol β, and epidermal growth factor receptor (EGFR) was evaluated using a 2-step quantitative reverse transcription polymerase chain reaction. Results: Fetal lung cell survival rate remained unaffected with 5-MTHF concentration below 1.25 µM. Beyond this concentration, cell viability is reduced with an increase in concentration. Cell cycle analysis revealed cell arrest in the G1 phase. The antioxidant activity of glutathione led the cells to bypass this arrest. Precisely, 10 and 50 µM 5-MTHF concentrations led to double-strand DNA breaks and single-strand DNA breaks. Gene expression study revealed lower expression of the MTR gene and higher expression of MTHFD1, EGFR, XRCC1, and DNA Pol β gene with an increase in 5-MTHF concentration. Conclusion: 5-MTHF concentration higher than 1.25 µM led to DNA damage in MRC5 and IMR90 human fetal lung cells.

CT-Scan-Assessed Body Composition and Its Association with Tumor Protein Expression in Endometrial Cancer: The Role of Muscle and Adiposity Quantities

Background: Endometrial cancer is strongly associated with obesity, and tumors often harbor mutations in major cancer signaling pathways. To inform the integration of body composition into targeted therapy paradigms, this hypothesis-generating study explores the association between muscle mass, body fat, and tumor proteomics. Methods: We analyzed data from 113 patients in The Cancer Genome Atlas (TCGA) and Cancer Proteomic Tumor Analysis Consortium (CPTAC) cohorts and their corresponding abdominal CT scans. Among these patients, tumor proteomics data were available for 45 patients, and 133 proteins were analyzed. Adiposity and muscle components were assessed at the L3 vertebral level on the CT scans. Patients were stratified into tertiles of muscle and fat mass and categorized into three groups: high muscle/low adiposity, high muscle/high adiposity, and low muscle/all adiposities. Linear and Cox regression models were adjusted for study cohort, stage, histology type, age, race, and ethnicity. Results: Compared with the high-muscle/low-adiposity group, both the high-muscle/high-adiposity (HR = 4.3, 95% CI = 1.0–29.0) and low-muscle (HR = 4.4, 95% CI = 1.3–14.9) groups experienced higher mortality. Low muscle was associated with higher expression of phospho-4EBP1(T37 and S65), phospho-GYS(S641) and phospho-MAPK(T202/Y204) but lower expression of ARID1A, CHK2, SYK, LCK, EEF2, CYCLIN B1, and FOXO3A. High muscle/high adiposity was associated with higher expression of phospho-4EBP1 (T37), phospho-GYS (S641), CHK1, PEA15, SMAD3, BAX, DJ1, GYS, PKM2, COMPLEX II Subunit 30, and phospho-P70S6K (T389) but with lower expression of CHK2, CRAF, MSH6, TUBERIN, PR, ERK2, beta-CATENIN, AKT, and S6. Conclusions: These findings demonstrate an association between body composition and proteins involved in key cancer signaling pathways, notably the PI3K/AKT/MTOR, MAPK/ERK, cell cycle regulation, DNA damage response, and mismatch repair pathways. These findings warrant further validation and assessment in relation to prognosis and outcomes in these patients.

MOLECULAR CLONING AND QUANTITATIVE mRNA EXPRESSION OF sox9 GENE IN GONADAL DEVELOPMENTAL PERIOD OF HYPOATHERINA TSURUGAE

Sox9 is a transcription factor of high mobility group (HMG) box family DNA binding domain. It plays a crucial role in gonadogenesis during embryonic developmental period. 1454 bp of sox9 mRNA transcript of Hypoatherina tsurugae (D. S. Jordan & Starks) was cloned and sequenced. It consists of an open reading frame (ORF) of 1436 bp, that encodes a 479 aa protein, found to be identical to the HMG box of other fish species. A phylogenetic tree was constructed by comparing the mRNA sequence of 50 different fishes across various taxa available in the NCBI database and using as outgroup Acipenser sinensis. The tree shows a high homology of sox9 from H. tsurugae with that from Maelanotaenia boesemani, the two forming a single clade. The expression of sox9 was studied in amhy+ (male) individuals. It begins from baseline at 0 wah (week after hatching) and is expressed in an increasing fashion. In amhy- (female) individuals it is highly expressed at initial stage (0 wah) and the expression reaches its peak at 2 wah then declines, indicating the low expression needed for differentiation of the female sex organs. The histological sections of gonads were studied in different stages of biweekly collected larvae during the sex determination/differentiation period and it showed that differentiation of gonads male/female is decided at 6 wah. In this stage the primary oocytes are clearly recognized and it correlates with the expression of sox9 genes. These finding add to the knowledge for a better understanding of molecular mechanisms of sex determination and differentiation period in fishes.

The expression and clinical significance of serum exosomal-long non-coding RNA DLEU1 in patients with cervical cancer

Background Accumulating evidence has demonstrated that the long non-coding RNA (lncRNA) lymphocytic leukaemia deletion gene 1 (DLEU1) is abnormally overexpressed in many cancer types, including cervical cancer (CC). However, the potential clinical significance of DLEU1 in serum exosomes of patients with CC remains unclear. Methods The expression of serum exosomal DLEU1 was detected by quantitative real-time polymerase chain reaction (qRT-PCR). A receiver operating characteristic (ROC) curve was plotted to evaluate the clinical diagnostic efficacy of DLEU1. The Kaplan–Meier survival curve and Cox proportional hazards model were used to assess the effect of DLEU1 on postoperative recurrence, metastasis and prognosis among patients with CC. Results Our research showed that DLEU1 expression in the serum exosomes of patients with CC was significantly upregulated compared to that in patients with cervical intraepithelial neoplasia (CIN) and healthy controls (HCs) (both p < .001). DLEU1 relative expression was significantly correlated with tumour size, cervical invasion depth, pathological grade, International Federation of Gynecology and Obstetrics (FIGO) stage and lymph node metastasis among patients with CC (p < .01 all). The combined detection of DLEU1, carbohydrate antigen 125 (CA-125) and squamous cell carcinoma (SCC) exhibited significantly higher diagnostic efficiency (p < .01). Furthermore, the overall survival (OS) and disease-free survival (DFS) of CC patients in the high DLEU1 expression group were markedly lower than those in the low DLEU1 expression group (both p < .01). Cox univariate and multivariate regression analyses indicated that DLEU1 was an independent risk factor for postoperative recurrence and metastasis in CC patients. Conclusions Our findings suggest that serum exosome DLEU1 has certain clinical value for diagnosing, monitoring recurrence and metastasis, and evaluating CC prognosis.

Identification of Gene‐Therapy Responsive Blood Biomarkers in mdx Mouse Model

ABSTRACTIntroductionIdentifying serum biomarkers that reflect the restoration of dystrophin in skeletal muscle is important for evaluating the effect of dystrophin‐restoring therapies in preclinical and clinical trials. Many potential blood biomarkers have been identified in Duchenne muscular dystrophy (DMD) patients, which change with disease progression or respond to pharmacological treatment. In this study, it was suggested that a panel of such blood biomarker candidates could be used to monitor dystrophin rescue in mdx mice treated with microdystrophin based therapies.MethodsPlasma samples from mdx mice treated with the microdystrophin therapy SGT‐001 were analysed with an antibody suspension bead array consisting of 87 antibodies. The array targets 83 unique proteins previously identified as biomarker candidates for DMD. Each sample was assayed at two different plasma dilutions to cover a broader concentration range. Protein concentrations estimated as Median fluorescent intensities (MFI) were correlated to dystrophin expression in muscle tissue, as measured by immunohistochemistry and Western blot. Thirteen of the targets were selected and analysed in a DMD and Becker muscular dystrophy (BMD) longitudinal natural history cohort using a suspension bead array.ResultsTen proteins were found to be significantly elevated in untreated mdx mice compared with C57 wild‐type mice and to correlate with dystrophin expression (Spearman's correlation, FDR &lt; 0.05) upon gene transfer in mdx mice. Translatability of these biomarkers from animal models to patients was evaluated by exploring abundance trajectories over time in both DMD and BMD patients and association with dystrophin expression in BMD patients. Consistent with the observations in mouse, six of these biomarker candidates were more abundant in DMD patients compared with controls, and six were also differentially abundant between BMD and DMD patients. Among them, serum titin was shown to be associated with dystrophin expression in BMD patients, having a steeper decline over time in patients with low dystrophin expression in tibialis anterior compared with patients with high expression. Myosine light chain 3 had a steeper decline with time in DMD patients compared with BMD patients.ConclusionsThe 10 biomarker candidates identified in mouse plasma are related to muscle contraction, glycolysis, microtubule formation and protein degradation. Human titin and myosine light chain 3 were the most interesting candidates as explorative biomarkers to monitor microdystrophin expression in gene therapies. If confirmed, these biomarkers could be used to detect restoration of dystrophin expression per se, monitor changes in dystrophin expression over time and potentially confirm disease phenotype changes from severe to mild disease forms.

Estimate the relationship between CXCR4-SDF-1 axis and inhibitory molecules (CTLA4 and PD-1) in patients with colon cancer

Colon neoplasia is one of the major malignancies in industrialized countries due to their Western-style food habits. It accounts for more than 50% of the population developing adenomatous polyps by the age of 70 years, but 10% of cancers in developed countries. The aim of this study was to evaluate the pathological role of the C-X-C chemokine receptor type 4/stromal-derived factor 1 axis (CXCR4-SDF-1 axis), and the inhibitory molecules PD-1 and cytotoxic T-lymphocyte associated protein 4 (CTLA-4) in postoperative colon cancer patients undergoing treatment with chemotherapy (oxaliplatin and capecitabine) and estimate the correlation between these studied factors to deeply understand the basic mechanisms and potential diagnostic or therapeutic effects. The study involved 90 patients, including 50 colon cancer patients (male and female, aged 35–65) diagnosed by oncologists at Al-Ramadi Hospital, Ramadi, Iraq. All patients underwent surgical resection and received four cycles of chemotherapy with oxaliplatin (85 mg every 21 days) and capecitabine (6 grams daily for 21 days). Additionally, 40 age- and sex-matched individuals served as the control group. For each participant, CXCR4 and SDF-1 levels were measured using ELISA and the gene expression of CTLA-4 and PD-1 were measured using RT-PCR. The colon cancer patient group showed significantly lower levels of CXCR4 and SDF-1 compared to control groups (0.163±0.012 vs 0.376±0.025 pg/mL and 0.376±0.025 vs 0.699±0.110 pg/mL, respectively, both had p=0.001). Moreover, the colon cancer patient group had significantly lower expression of PD-1 and CTLA4 compared to control group (0.102±0.029-fold vs 1.199±0.391-fold, p=0.02; and 0.302±0.140-fold vs 1.441±0.334-fold, p=0.008, respectively). In conclusion, the results suggest that CXCR4 and SDF-1 appear promising as diagnostic markers for distinguishing colon cancer patients from healthy conditions.

Gastric Adenocarcinomas with CDX2 Induction Show Higher Frequency of TP53 and KMT2B Mutations and MYC Amplifications but Similar Survival Compared with Cancers with No CDX2 Induction

Background: Gastric cancer is one of the most prevalent gastrointestinal cancers. Mortality is high, and improved treatments are needed. A better understanding of the pathophysiology of the disease and discovery of biomarkers for targeted therapies are paramount for therapeutic progress. CDX2, a transcription factor of hindgut specification, is induced in several gastric cancers, especially with intestinal differentiation, and could be helpful for defining sub-types with particular characteristics. Methods: Gastric cancers with induced CDX2 mRNA expression were identified from the gastric cohort of The Cancer Genome Atlas (TCGA) and were compared with cancers that had no CDX2 mRNA induction. Induced CDX2 mRNA expression was defined as mRNA expression z-score relative to all samples above 0, and non-induced CDX2 mRNA expression was defined as mRNA expression z-score relative to all samples below −1. Results: Patients with gastric cancers with CDX2 mRNA induction were older, had less frequently diffuse histology, and more often had mutations in TP53 and KMT2B and amplifications in MYC. CDX2 induction was correlated with HNF4α induction and was reversely correlated with SOX2. Gastric cancers with CDX2 mRNA induction showed lower PD-L1 expression than cancers with lower CDX2 expression but did not differ in CLDN18 mRNA expression. Progression-free and overall survival of the two groups was also not significantly different. Conclusion: Gastric cancers with CDX2 mRNA induction displayed specific characteristics that differentiate them from cancers with no CDX2 induction and could be of interest for optimizing current and future therapies.

Matrine promotes colorectal cancer apoptosis by downregulating shank-associated RH domain interactor expression

BACKGROUND The 5-year survival rate of patients with colorectal cancer (CRC) in China is only 56.9%, highlighting the need for new therapeutic drugs. Previous studies have shown that matrine exhibits antitumor effects by inducing apoptosis. However, the mechanism by which matrine regulates antiapoptotic proteins in CRC remains unclear. AIM To identify apoptotic proteins from proteomics and investigate the role of matrine in impeding CRC apoptosis by regulating these proteins. METHODS Tumor and adjacent normal tissues were collected from 52 patients with CRC who underwent surgery between January and December 2021. Data-independent acquisition quantitative proteomic analysis was performed to identify differentially expressed apoptotic proteins. The selected apoptotic proteins were identified through their association with tumor-node-metastasis (TNM) stage and prognosis, then confirmed by immunohistochemical (IHC) staining in validation cohort. In vitro , the role of matrine or apoptotic proteins on cancer cells were analyzed. RESULTS Compared to normal tissues, 88 anti-apoptotic proteins from proteomic results were selected. Among them, Shank-associated RH domain interactor (SHARPIN) was identified because of its relationship with TNM stage and overall survival in TCGA database. In the IHC-confirmed cohort, SHARPIN was highly expressed in CRC tissues and localized in the cytoplasm. Higher SHARPIN expression was associated with TNM stage, carbohydrate antigen 153 levels, and gross type compared to low expression. SHARPIN knockdown promoted apoptosis, significantly upregulated the expression of Bcl-2 associated agonist of cell death, Bcl-2 associated X protein, caspase 3, and caspase 8, and downregulated B-cell lymphoma-2 (P &lt; 0.05). Importantly, matrine treatment promoted apoptosis and reversed the proliferation, invasion, and migration of CRC cells by repressing SHARPIN. CONCLUSION SHARPIN was identified as an upregulated anti-apoptotic protein in CRC, and matrine exhibited anticancer effects by downregulating its expression. Thus, matrine appears to be a promising drug for CRC.

The Expression of SIRT3 in Endometrial Carcinoma and Its Effect on Promoting Apoptosis of Ishikawa Cells.

Endometrial cancer (EC) is one of the three most common malignancies of the female reproductive system. SIRT3 is an NAD+-dependent protein deacetylase that maintains the stability of the intracellular environment. This study aims to investigate the mechanism of SIRT3 in regulating apoptosis in endometrial cancer and further reveal the role of SIRT3 in endometrial cancer. Differential expression of SIRT3 in tumors was analyzed by GEPIA using TCGA database data. Meanwhile, mRNA and protein expression levels of SIRT3 in tissues and cells were examined using RT-qPCR, Western Blot, and immunohistochemistry. The expression of SIRT3 after estradiol (E2) stimulation of Ishikawa cells was detected using RT-qPCR and Western Blot techniques. The effect of transfection after SIRT3 knockdown and overexpression was verified using RT-qPCR and Western Blot. Flow cytometry and TUNEL assay were used to detect the effect of SIRT3 on apoptosis. Reactive oxygen species (ROS) was used to detect the effect of SIRT3 on the level of oxidative stress in cells. The expression of apoptotic protein (BAX, cleaved-Caspase 3) and autophagy protein (cyto C and LC3A) were detected in transfected Ishikawa cell. Differences analysis of TCGA database data showed that the expression of SIRT3 in EC was significantly lower than that in normal endometrial tissue. The mRNA and protein levels of SIRT3 were significantly lower in EC tissues or cells than normal controls. E2 stimulation in Ishikawa cells resulted in the down-regulation of SIRT3 expression. After transfection, SIRT3 promoted the apoptosis of Ishikawa cells and attenuated the levels of ROS. Overexpression of SIRT3 promoted apoptosis and autophagy-related proteins. Thus, high expression of SIRT3 inhibits the development of EC whereas low expression of SIRT3 may promote the progression of EC, which provides a new direction for studying the treatment of EC.

Elevated IL-6 Expression in Autologous Adipose-Derived Stem Cells Regulates RANKL Mediated Inflammation in Osteoarthritis

Interleukin-6 (IL-6) expression in mesenchymal stem cells (MSCs) has been shown to play a pivotal role in modulating cartilage regeneration and immune responses, particularly in the context of diseases that involve both degenerative processes and inflammation, such as osteoarthritis (OA). However, the precise mechanism through which IL-6 and other immune-regulatory factors influence the therapeutic efficacy of autologous adipose-derived stem cells (ASCs) transplantation in OA treatment remains to be fully elucidated. This study aims to investigate the relationship between IL-6 expression in autologous ASCs isolated from OA patients and their impact on immune modulation, particularly focusing on the regulation of Receptor Activator of Nuclear factor Kappa-Β Ligand (RANKL), a key mediator of immune-driven cartilage degradation in OA. Autologous ASCs were isolated from the stromal vascular fraction (SVF) of adipose tissue obtained from 22 OA patients. The isolated ASCs were cultured and characterized using reverse transcription polymerase chain reaction (RT-PCR), enzyme-linked immunosorbent assay (ELISA), and flow cytometry to the phenotype and immune regulatory factors of MSCs. Based on IL-6 expression levels, ASCs were divided into high and low IL-6 expression groups. These groups were then co-cultured with activated peripheral blood mononuclear cells (PBMCs) to evaluate their immune-modulatory capacity, including the induction of regulatory T cells, inhibition of immune cell proliferation, and regulation of key cytokines, such as interferon-gamma (IFN-γ). Additionally, RANKL expression, a critical factor in osteoclastogenesis and cartilage degradation, was assessed in both ASC groups. High IL-6-expressing ASCs demonstrated a significantly greater capacity to inhibit immune cell proliferation and IFN-γ production compared to their low IL-6-expressing counterparts under co-culture conditions. Moreover, the group of ASCs with high IL-6 expression showed a marked reduction in RANKL expression, suggesting enhanced potential to control osteoclast activity and subsequent cartilage defect in OA. Conclusion: Autologous ASCs with elevated IL-6 expression exhibit enhanced immunomodulatory properties, particularly in regulating over-activated immune response and reducing osteoclastogenesis through RANKL suppression. These findings indicate that selecting ASCs based on IL-6 expression could enhance the therapeutic efficacy of ASC-based treatments for OA by mitigating immune-driven joint inflammation and cartilage degradation, potentially slowing disease progression.