Protein Levels In Tissues Research Articles

Unveiling Molecular Dynamics of MeCp2, CDKL5 and BDNF in the Hippocampus of Individuals With Intractable Mesial Temporal Lobe Epilepsy.

Mutations occurring in the MeCp2, CDKL5 and BDNF genes have been linked to epileptogenesis in various epilepsy syndromes. This study employed bioinformatics analysis of transcriptomic data to examine the interrelationship among these genes in both epileptic and healthy individuals. Moreover, we assessed the expression of MeCp2, CDKL5 and BDNF at both mRNA and protein levels in human hippocampal tissues obtained from 22 patients undergoing epilepsy surgery for mesial temporal lobe epilepsy (MTLE) as well as from 25 autopsied specimens. Bioinformatics findings suggest that MeCp2, CDKL5 and BDNF genes play a role in regulating genes associated with epilepsy and disruptions in these genes may contribute to epilepsy development. Furthermore, the study reveals significantly lower MeCp2 and CDKL5 protein levels in the epileptic hippocampus compared to controls. Positive correlations are observed between MeCp2 and CDKL5 mRNA expression in autopsied samples and between CDKL5 and BDNF mRNA expression in epileptic hippocampal tissues. Differences in mRNA expression correlation patterns of MeCp2 and CDKL5 with BDNF are found between epileptic and control hippocampal tissues. Moreover, a significant positive correlation between MeCp2 and CDKL5 protein expression is noted in control hippocampal tissues. Our data suggest that altered expression of MeCp2, CDKL5 and BDNF within the hippocampus may contribute to epileptogenic processes in MTLE, impacting seizure characteristics, surgical outcomes and responses to antiepileptic drugs. Alterations in the expression of MeCp2, CDKL5 and BDNF within the hippocampus might contribute to the epileptogenic processes in MTLE. These changes could be linked to distinct functional consequences in epilepsy.

Study on the role and mechanism of TMED2 in oral squamous cell carcinoma.

This study investigated TMED2 expression in oral squamous cell carcinoma (OSCC) and its effects on SCC9 cell behaviors, including proliferation, migration, invasion, and autophagy, to support OSCC diagnosis and treatment. TMED2 expression was analyzed in TCGA and GEO databases, and protein levels in OSCC tissues were examined via HE staining and tissue microarrays. SCC9 cells, with high TMED2 expression, were used to assess TMED2's effects on cell proliferation, invasion, and cell cycle. TMED2 knockdown was performed with lentiviral vectors, and RT-PCR and Western blotting explored the autophagy and AKT/mTOR pathways. Tumor growth was tested in TMED2 knockdown and control cells in nude mice. TMED2 was highly expressed in OSCC, correlating with poor prognosis. Knockdown of TMED2 significantly reduced SCC9 cell proliferation, migration, and invasion, induced G0/G1 cell cycle arrest, reduced AKT/mTOR pathway activity, and increased autophagy, prolonging survival in tumor-bearing mice. TMED2 is upregulated in OSCC, correlates with poor prognosis, and regulates cell proliferation, invasion, and autophagy, indicating it as a potential therapeutic target.

PRDX6 Prevents NNMT Ubiquitination and Degradation as a Nonenzymatic Mechanism to Promote Ovarian Cancer Progression.

Cancer cells cope with oxidative stress for their proliferation and metastasis by equipping antioxidant systems, among which the antioxidant enzymes peroxiredoxins (PRDXs) play crucial roles. However, whether PRDXs exhibit nonenzymatic functions remains unclear. Here, it is shown that the 1-cysteine PRDX (PRDX6) upregulates nicotinamide N-methyltransferase (NNMT) to promote the growth and metastasis of ovarian cancer cells, independently of PRDX6's enzymatic activities. Mechanistically, PRDX6 interacts with NNMT to prevent its binding to the E3 ubiquitin ligase tripartite-motif protein 56 (TRIM56), leading to the inhibition of NNMT ubiquitination at lysine 23 and 210 and suppression of subsequent proteasomal degradation. In addition, PRDX6-mediated NNMT upregulation activates mitogen-activated protein kinase (MAPK) signaling, thereby promoting the growth and metastasis of ovarian cancer cells. Notably, PRDX6 overexpression is associated with higher NNMT protein levels in human ovarian cancer tissues and is predictive of poor prognosis of ovarian cancer patients. Overall, the findings illustrate a critical oncogenic mechanism of the antioxidant enzyme PRDX6 in promoting ovarian cancer progression beyond its enzymatic mechanisms.

Expression of fatty acid binding proteins in mesenteric adipose tissue.

Adipose is a complex tissue comprised of adipocytes, immune cells, endothelial and progenitor stem cells. In humans, there are at least nine defined adipose depots, each containing variable numbers of genetically identified adipocyte clusters suggesting remarkable heterogeneity and potential functionality in each depot with respect to lipid metabolism. Although subcutaneous and visceral depots are commonly analyzed for biochemical and molecular functions, the mesenteric depot has been overlooked yet strongly implicated in lipid mediated immune surveillance. Since fatty acid binding proteins (FABPs) are primary cellular conduits to lipid trafficking, we evaluated the expression patterns for four major fatty acid binding proteins (FABP1, FABP3, FABP4 and FABP5) using a combination of gene expression, immunoblotting, and immunofluorescence in mesenteric fat from both young and old, male and female C57Bl/6J mice. All four FABPs were expressed at the mRNA and protein level in murine mesenteric adipose tissue. While there was no statistical change in expression of mesenteric FABP isoforms with sex or age, the expression of mesenteric FABP1 was increased, and FABP4 decreased, in both males and females as compared to perigonadal and inguinal depots. Surprisingly, immunofluorescence staining revealed that compared to subcutaneous or perigonadal depots, mesenteric fat expresses FABP3, but little FABP5, in adipocytes. These results highlight the diversity in adipose tissue and the importance of evaluating the mesenteric depot in the context of lipid transport and metabolism.

HMGB2 knockdown ameliorates retinal ganglion cell injury by inhibiting NLRP3 inflammasome activation after retinal ischemia.

To explore the neuroprotective effects of high mobility group box 2 (HMGB2) knockdown on retinal ganglion cells (RGCs) in the retinal ischemia-reperfusion injury (RIRI). Oxygen-glucose deprivation (OGD)-injured RGCs from postnatal three-day C57BL/6 mice pups and high intraocular pressure (IOP)-induced RIRI mice were used as cellular and animal models of RIRI. The expression of HMGB2 in the retina of RIRI mice and OGD-injured RGCs was detected through reverse transcription-polymerase chain reaction (RT-qPCR) and Western blotting. The effects of HMGB2 silencing on the morphological changes, RGCs survival, and cell apoptosis in mouse retinal tissues were observed through H&E staining, immunofluorescence staining with RNA-binding protein with multiple splicing (RBPMS) antibody, and TUNEL staining, respectively. RGC viability and apoptosis were examined by CCK-8 and flow cytometry assays. The levels of proteins associated with NOD-like receptor thermal protein domain associated protein 3 (NLRP3)-mediated pyroptosis [NLRP3, Caspase-1, GSDMD-N, interleukin (IL)-1β, IL-18] in vivo and in vitro were measured by Western blotting. HMGB2 protein and NLRP3 were upregulated in the retina of RIRI mice and OGD-injured RGCs (P<0.001). The retina was edematous, accompanied by disorganized cell arrangement and decreased thickness of all layers, and obvious vacuoles in ganglion cell layer. HMGB2 silencing alleviated the reduction in total retinal thickness and the severity of retinal tissue damage as well as suppressed RGC loss and retinal cell apoptosis in RIRI mice. OGD-induced RGC apoptosis was ameliorated after downregulation of HMGB2 in vitro. Intravitreal injection of the AAV-sh-HMGB2 and si-HMGB2 resulted in significantly decrease of NLRP3, Caspase-1, GSDMD-N, IL-1β, and IL-18 protein levels in the retinal tissues of RIRI mice and OGD-injured RGCs, respectively (all P<0.001). HMGB2 knockdown protects against RGC apoptosis and pyroptosis after RIRI through suppressing NLRP3 inflammasome activation.

Investigation of the Mesencephalic Astrocyte Derived Neurotrophic Factor-Endoplasmic Reticulum Stress Pathway in Mood Disorders.

Bipolar disorder (BD) has been associated with impaired cellular resilience. Recent studies have shown abnormalities in the unfolded protein response (UPR) in BD. The UPR is the cellular response to endoplasmic reticulum (ER) stress. Mesencephalic astrocyte-derived neurotrophic factor (MANF), a trophic factor, decreases ER stress by modulating the UPR. The objective of this study is to investigate the MANF-ER stress pathway in BD and major depressive disorder (MDD) compared to healthy controls (HC). MANF protein concentration and MANF and GRP78 gene expression were assessed in peripheral blood from individuals with BD, MDD and HC (protein: 40 BD, 55 MDD, 55 HC; gene expression: 52 BD, 61 MDD, 69 HC). MANF protein and gene expression along with GRP78 gene expression were also analyzed in postmortem brain tissue (20 BD, 20 MDD, 19 HC). MANF protein was quantified using an ELISA assay while quantitative polymerase chain reaction was used for MANF and GRP78 gene expression. Peripheral MANF protein levels were reduced in individuals with BD in a depressive state compared to controls (p=0.031) and euthymic BD participants (p=0.013). No significant differences in MANF or GRP78 gene expression were observed in BD irrespective of mood state, or MDD compared to HC (all p>0.05). No differences were observed regarding MANF/GRP78 protein or gene expression levels in postmortem tissue (p>0.05). Individuals with BD who were in an acute depressive phase were found to have reduced peripheral MANF levels potentially signifying abnormal UPR and supporting the notion that BD is associated with increased ER stress.

Best1 mitigates ER stress induced by the increased cellular microenvironment stiffness in epilepsy.

Changes in brain tissue stiffness are closely linked to the development and diseases of the nervous system. Endoplasmic reticulum (ER) stress plays a role in various pathological processes related to epilepsy. However, the relationship between stiffness changes, ER stress, and epilepsy remains unclear. This study aimed to investigate the impact of Best1 upregulation on alleviating ER stress and the underlying mechanism. Additionally, we proposed a protective strategy to prevent cell death resulting from ER stress in epilepsy. This study investigated the expression levels of ER stress-related proteins in epileptic tissues of varying stiffness. Atomic force microscopy revealed differences in stiffness across various lesion regions in patients with epilepsy. The expression levels of ECM and ER stress-related proteins were elevated in tissues with higher stiffness. Polypropionamide hydrogels were used to simulate extracellular matrix (ECM) with varying stiffness levels. Basal ER stress increased in the stiffer hydrogel substrates. Furthermore, the calcium-activated anion channel Bestrophin 1 (Best1) mitigated ER stress induced by both the stiffer substrate and thapsigargin. The loss-of-function mutations in Best1 inhibited this activity. The underlying mechanism involves the upregulation of the endosomal sorting complex required for the transport (ESCRT) components by Best1, which helps mitigate ER stress. These findings suggest that increased stiffness of the cellular microenvironment may contribute to neuronal death during epileptogenesis. Additionally, Best1 upregulation may serve as a protective strategy against excessive ER stress-induced neuronal damage in epilepsy.

Ellagic acid alleviates NLRP6/caspase-1/GSDMD-mediated inflammation and pyroptosis in rats post cerebral ischemia/reperfusion injury.

Ellagic acid (EA) is a natural polyphenol with anti-cancer, anti-oxidant, anti-inflammatory, antibacterial, and other effects. However, the role of EA in cerebral ischemia/reperfusion injury (CIRI) remains unclear. This study aims to investigate the neuroprotective effects of EA in CIRI. Forty male Wistar rats (260-300 g) were randomly divided into four groups with 10 rats per group: 1) Sham+Veh: Rats underwent I/R surgery, except that they were not inserted with thread plugs, and received solute treatment at the same time. 2) MCAO/R+Veh. 3) MCAO/R+EA: Rats were administered 200 mg/kg EA before undergoing MCAO. 4) MCAO/R+Nim: Rats were administered Nim before undergoing MCAO. Cerebral MCAO/R damaged brain tissue, elevated neurological deficit score (P<0.01), cerebral infarction volume (P<0.01), inflammatory cell infiltration (P<0.01), NLRP6, ASC, caspase-1 and GSDMD mRNA level (P<0.01 and P<0.001), NLRP6, caspase-1, GSDMD-N and IL-1β protein level (P<0.01 and P<0.001), and inflammatory cytokines in brain tissue (P<0.01). Prophylactic administration of EA also significantly improved brain tissue damage, reduced neurological deficit score (P<0.01), cerebral infarction volume (P<0.01), inflammatory cell number (P<0.05), NLRP6, caspase-1, GSDMD-N mRNA and protein level (P<0.05 and P<0.01), ASC mRNA level and IL-1β protein level (P<0.01), and IL-1β and IL-18 level in brain tissue (P<0.01) compared to positive control. EA may serve as a potential drug for the treatment of brain I/R, which may exert an anti-inflammatory effect by inhibiting the activation of the inflammasome.

Expression of lipid metabolism and cartilage degeneration-related factors in lumbar vertebral endplate Modic changes.

This study aims to investigate the relationship between the expression of lipid metabolism and cartilage degeneration-related factors and Modic changes (MCs) of lumbar vertebral. This prospective study included a total of 10 patients (6 males, 4 females; mean age: 60.4±8.7 years; range 51 to 82 years) who underwent lumbar interbody fusion surgery due to degenerative lumbar diseases (MC group), and 10 control patients (4 males, 6 females; mean age: 49.7±9.8 years; range, 42 to 76 years) with lumbar burst fractures (nonMC group) between January 2020 and December 2022. Clinical imaging data and cartilage tissues were collected to observe cartilage characteristics and pathological changes. The relative expression levels of lipid metabolism-related inflammatory factors matrix metalloproteinase-1 (MMP-1), a disintegrin and metalloproteinase with thromboSpondin motifs-5 (ADAMTS-5), and aggrecan in cartilage were detected by quantitative polymerase chain reaction (qPCR). The relative expression levels of MMP-1 and ADAMTS-5 proteins in cartilage tissues were detected by Western blotting. There were no significant differences in the baseline characteristics between the two groups. The color and transparency of the endplate cartilage in the control group were significantly better than those in the MCs group. Radiographic and hematoxylin-eosin staining of the endplate cartilage tissues showed that the extracellular matrix was higher in the control group than in the MCs group (p<0.05). Compared to the control group, qPCR analysis showed higher expression of MMP-1 and ADAMTS-5 in the MCs group, while aggrecan expression was lower (p<0.05). Western blot analysis showed that both MMP-1 and ADAMTS-5 expression were higher in the MCs group than in the control group (p<0.05). Lipid metabolism and cartilage degeneration-related inflammatory factors exist in the vertebral endplate of the patients with degenerative lumbar diseases, and the upregulation of MMP-1 and ADAMTS-5 may be related to MCs and endplate degeneration.

Integrated lipidomics and RNA-seq reveal prognostic biomarkers in well-differentiated and dedifferentiated retroperitoneal liposarcoma

BackgroundRetroperitoneal liposarcoma (RLPS) is a mesenchymal malignant tumor characterized by different degrees of adipocytic differentiation. Well-differentiated liposarcoma (WDLPS) and dedifferentiated liposarcoma (DDLPS) are two of the most common subtypes of RLPS, exhibiting clear differences in biological behaviors and clinical prognosis. The metabolic features and genomic characteristics remain unclear.MethodsThis study employed lipidomic and RNA-seq analyses of RLPS tissues from 19 WDLPS and 29 DDLPS patients. Western blot and immunohistochemistry staining were performed to verify the tumor tissue protein levels of TIMP1, FN1, MMP11, GPNMB, and ECM1. Enzyme-linked immunosorbent assay (ELISA) was performed to evaluate different serum protein levels in 128 blood samples from patients with RLPS. Multivariate analysis was performed to identify the most crucial variables associated with overall survival (OS) and recurrence-free survival (RFS) of the RLPS patients.ResultsLipidomic analysis revealed a significant difference in lipid metabolism, particularly in phosphatidylcholines and triacylglycerides metabolism. RNA sequencing analysis revealed that 1,630 differentially expressed genes (DEGs) were significantly enriched in lipid metabolism, developmental process, and extracellular matrix (ECM) pathways. Integrated lipidomic and transcriptomic analysis identified 29 genes as potential biomarkers between WDLPS and DDLPS. Among the 29 DEGs, we found that TIMP1, FN1, MMP11, GPNMB, and ECM1 were increased in DDLPS tumor tissues than in WDLPS tumor tissues. The receiver operating characteristic (ROC) curve showed high specificity and sensitivity in diagnosing patients using a five-gene combination (AUC = 0.904). ELISA revealed a significant increase in the serum levels of ECM1 and GPNMB in patients with DDLPS compared to patients with WDLPS. ECM1 increased progressively across different FNCLCC Grades, correlating negatively with RFS (P = 0.043). GPNMB levels showed a negative correlation with OS (P = 0.019).ConclusionsOur study reveals different lipid metabolism, several transcriptional pathways between WDLPS and DDLPS, and examines several serum markers associated with the prognosis of RLPS. These findings provide a vital basis for future endeavors in diagnosing and predicting the prognosis of retroperitoneal liposarcoma with different differentiations.

Preliminary Study on the Positive Expression Regulation of Alpha2-Macroglobulin in the Testicular Tissue of Male Mice by Environmental Estrogens.

The male reproductive impairment caused by environmental estrogens (EEs) stands as a pivotal research area in environmental toxicology. Alpha2-macroglobulin (A2M) emerges as a promising molecule capable of counteracting oxidative stress induced by EEs. This study conducted exposure experiments spanning PND1 to PND56 employing ICR mice, aiming to delve into the expression patterns of A2M and its modulated IL-6 in the testicular tissue of mice subsequent to diethylstilbestrol (DES) and benzophenone (BP) exposure, while elucidating the pivotal role of ERs in this intricate process. Our findings revealed that upon DES exposure (10 and 100 nM), there was a pronounced upregulation of A2M (mRNA and in situ protein levels) in mouse testicular tissue. Similarly, exposure to BPs (BP-1, BP-2, and BP-3, each at 10 and 1000 nM) exhibited comparable effects and increasing A2M levels in serum. Notably, BP exposure also caused an elevation in IL-6 levels (which could be directly regulated by A2M) within testicular tissue (mRNA and in situ protein). Remarkably, the specific estrogen receptor antagonist ICI 182780 (0.5 mg/kg/day) was effective in reversing the upregulation of both A2M and IL-6 induced by BP exposure. Significantly, the results of theoretical prediction of the potential ERE site in the A2m gene promoter region and ChIP-qPCR experiment provide essential and strong evidence for the key conclusion that A2m is the target gene of ER. Taken together, our study highlights EEs' ability to regulate A2M expression in the male reproductive system via the ER signaling pathway. This vital insight deepens our understanding of molecular mechanisms protecting against oxidative stress caused by EEs.

Effects of ginger root extract on cognitive impairment and the Nrf2/GPX4/SLC7A11 signaling pathway in aluminum-exposed rats.

Aluminum exposure has been widely demonstrated to cause cognitive impairment and neuronal damage, which are significant factors in many neurodegenerative diseases. Ginger root extract (GRE) has attracted researchers' attention in recent years due to its antioxidant and neuroprotective properties, as a potential therapeutic intervention to mitigate aluminum-induced neural damage.To evaluate the effects of GRE on cognitive impairment and the expression of Nrf2, GPX4, and SLC7A11 in aluminum (Al)-exposed rats. In this study, a total of 60 male Sprague-Dawley (SD) rats were randomly assigned to 6 groups: Control, Al exposure (Al), Clinical Drug, low-dose GRE (Al + L-GRE), medium-dose GRE (Al + M-GRE), and high-dose GRE (Al + H-GRE). All groups, except Control, received 10mg/ml AlCl₃ in drinking water for three months. Post-exposure treatments included saline for Control and Al groups, 0.5mg/kg donepezil hydrochloride for the Clinical Drug group, and 100, 200, and 400mg/kg GRE for the respective GRE groups. Treatments were administered once daily, five days a week, for a duration of one month. Cognitive and spatial abilities were assessed using the Morris water maze and Y-maze tests. Neuronal damage and mitochondrial structure in the hippocampus were evaluated via Nissl staining and transmission electron microscopy. Serum levels of SOD, GSH, MDA, and Fe²⁺ were measured, and Nrf2/GPX4/SLC7A11 mRNA and protein levels in hippocampal tissues were analyzed using real-time PCR and Western blotting. The Al group demonstrated compromised cognitive function, diminished neuronal integrity, disrupted mitochondrial structure, and an imbalance in serum antioxidant levels compared to the Control group.GRE treatment appeared to have a positive effect on cognitive function, potentially contributing to the maintenance of neuronal integrity, possibly aiding in the preservation of mitochondrial structure, and potentially assisting in the regulation of antioxidant levels. Notably, the Al + M-GRE and Al + H-GRE groups demonstrated encouraging enhancements in escape latency, platform crossings, novel arm entries, and time spent in the novel arm. Additionally, these groups showed elevated levels of SOD and GSH, reduced levels of MDA and Fe²⁺, and increased expression of Nrf2, GPX4, and SLC7A11, suggesting a potential beneficial impact of GRE treatment.GRE mitigated cognitive impairment in Al-exposed rats, likely through the Nrf2/GPX4/SLC7A11 signaling pathway, demonstrating its potential neuroprotective effects.

Villin-1 regulates ferroptosis in colorectal cancer progression.

Colorectal cancer (CRC) is the second leading cause of cancer-related deaths worldwide. Despite extensive research, the mechanistic underpinnings driving CRC progression remain largely unknown. As a fundamental component of the brush border cytoskeleton, villin-1 (VIL1) acts as a marker for intestinal cell differentiation and maturation. Through a comprehensive transcriptomics analysis of eight studies (total sample: n = 1952), we consistently observed significant upregulation of VIL1 expression in CRC tumors compared with adjacent normal tissue. In our independent cohort, this notable upregulation has been further validated at both mRNA and protein levels in colon tumor tissues, relative not only to adjacent normal tissue but also to normal controls. Our data show that VIL1 promotes proliferation and migration while inhibiting apoptosis. Conversely, knockout of VIL1 suppresses proliferation and migration while inducing apoptosis. Mechanistically, we reveal that knocking out VIL1 activates ferroptosis and inhibits the migration of CRC cells, while overexpressing VIL1 yields the opposite effects, and vice versa. Additionally, VIL1 binds to Nuclear factor NF-kappa-B p105 subunit (NF-κB) and controls NF-κB expression. In vivo, overexpressing VIL1 inhibits ferroptosis, and induces the expression of NF-κB and lipocalin 2 (LCN2), thereby promoting CRC tumor growth. Thus, we have identified the VIL1/NF-κB axis as a pivotal regulator of CRC progression through ferroptosis modulation, unveiling VIL1 as a promising therapeutic target for CRC treatment via ferroptosis. Our study offers novel avenues for exploring the therapeutic potential of ferroptosis in CRC management, emphasizing the high potential of VIL1 in regulating colorectal tumorigenesis.

KDM1A epigenetically enhances RAD51 expression to suppress the STING-associated anti-tumor immunity in esophageal squamous cell carcinoma

Histone lysine demethylase LSD1, also known as KDM1A, has been found to regulate multiple cancer hallmarks since it was first identified in 2004. Recently, it has emerged as a promising target for stimulating anti-tumor immunity, specifically boosting T cell activity. However, it remains unclear whether and how it remodels the tumor microenvironment to drive oncogenic processes in esophageal squamous cell carcinoma (ESCC). In this study, protein levels in ESCC tissues were evaluated by immunostaining of tissue microarrays. Cell growth was assessed by colony formation assays in vitro and subcutaneous xenograft models in vivo. High-throughput transcriptomics and spatial immune proteomics were performed using bulk RNA sequencing and digital spatial profiling techniques, respectively. Epigenetic regulation of RAD51 by methylated histone proteins was analyzed using chromatin immunoprecipitated quantitative PCR assays. Finally, our clinical data indicate that KDM1A precisely predicts the overall survival of patients with early-stage ESCC. Inhibition of KDM1A blocked the growth of ESCC cells in vitro and in vivo. Mechanistically, our transcriptomics and spatial immune proteomics data, together with rescue assays, demonstrated that KDM1A specifically removes methyl residues from the histone protein H3K9me2, a transcription repressive marker, thus reducing its enrichment at the promoter of RAD51 to epigenetically reactivate its transcription. Additionally, it significantly inhibits the expression of NF-κB signaling-dependent proinflammatory genes IL-6 and IL-1B through RAD51, thus blocking the STING-associated anti-tumor immunity in stromal tumor-infiltrating lymphocytes (sTIL). Overall, our findings not only indicate that KDM1A is a promising target for ESCC patients at early stages but also provide novel mechanistic insights into its spatial regulation of STING-associated anti-tumor immunity in sTILs to drive the oncogenic processes in ESCC. The translation of these findings will ultimately guide more appropriate combinations of spatial immunotherapies with KDM1A inhibitors to improve the overall survival of specific subgroups in ESCC.

Evaluation of Breast Cancer Gene Type 1 (BRCA1) Protein Levels in Cancer Tissue Using Surface-Enhanced Raman Spectroscopy.

Raman spectroscopy is a chemical process that utilizes the interaction between light and matter to get significant insights into the structure or characteristics of matter. Raman spectroscopy techniques, such as quantitative evaluation, early diagnostic capabilities, and elucidation of the spectral properties of tissues, are excellent candidates for use in research. In cancer, changes in genes and proteins expressed by related genes are associated with a poor prognosis and aggressive tumor characteristics. Due to modifications and regulatory steps in protein translation, the results of the messenger RNA (mRNA) expression of genes may not correctly reflect the results of protein expression. For this reason, the mRNA and protein expressions of genes are studied in parallel in molecular studies on cancer. In our study, the breast cancer gene type 1 (BRCA1) gene, which is frequently studied in breast cancer and is relatively more difficult to measure by traditional methods due to its high molecular weight, was selected, and protein quantification was performed in tissue samples by Raman spectroscopy. With Raman spectroscopy, it is possible to obtain rapid and precise quantitative results even with a small amount of sample, so it is quite advantageous compared to traditional methods. In our study, we performed surface-enhanced Raman spectroscopy (SERS) to analyze the quantitative protein amount. SERS is a highly sensitive method for detecting compounds at low concentrations. For this purpose, magnetic nanoparticles modified with protein antibodies were used, and the target protein was withdrawn from the complex environment and transferred to an appropriate buffer environment. The calibration curve for BRCA1, which plots Raman intensity against concentration, was derived by calculating the average response reading from duplicate assays conducted under identical conditions. The BRCA1 protein levels of cells were determined from the regression curve of the BRCA1 protein. The relation between the concentration of BRCA1 protein and SERS spectrum intensity was determined to be logarithmic in the range of 300 µg·mL-1 to 292 ng·mL-1 (R2 = 0.9928, limit of detection = 10.41 µg·mL-1, and limit of quantitation = 31.24 µg·mL-1).

Increased levels of DNA methyltransferases in the placentas of women affected by Placenta Accreta Spectrum lead to aberrant DNA methylation patterns.

Placenta Accreta Spectrum (PAS) is a serious placental abnormality assosiated with significant maternal death during pregnancy. Due to its invasive characteristics resembling tumor growth, PAS is often associated with tumor-related proteins. While DNA methylation is known to regulate genes involved in development, its potential role in the development of PAS remains unclear. The aim of our study was to investigate the impact of PAS on DNA methylation and DNA methyltransferases (DNMTs). The groups were categorized into three groups: vaginal birth, cesarean delivery, and cesarean delivery with PAS. To measure DNMT protein levels in placental tissue, we employed immunohistochemistry (IHC) and Western blot (WB) techniques. Global DNA methylation levels were assessed using 5-methylcytosine staining. We found that DNMT1, DNMT3A and DNMT3L levels were significantly increased in the placentas of PAS group compared to control counterparts in both WB and IHC analysis. Compatible with DNMTs expressions, global DNA methylation levels were found to be significantly higher in placentas from women with PAS compared to controls. Our results suggest that significant alterations in DNMTs expressions and global DNA methylation within placentas may contribute to the development of Placenta Accreta Spectrum (PAS). To elucidate the precise molecular mechanisms underlying these changes and their role in PAS pathogenesis, further research required.

Revisiting the follicle-stimulation hormone receptor expression and function in human myometrium and adipose tissue

BackgroundExtragonadal follicle-stimulating hormone receptor (FSHR) expression at low levels has been shown in several normal and tumor tissues, including myometrium and adipose tissue. FSH-FSHR signaling in the myometrium has been suggested to regulate uterine contractile activity and the timing of labor. In contrast, FSH-FSHR has been linked to the activation of brown/beige fat thermogenesis in adipose tissue. The issue of extragonadal FSHR expression and its functionality remains contentious within the scientific community, as contradictory findings necessitate further independent and critical analyses. Hereby, we re-investigated the FSHR expression and its functionality in normal non-pregnant (M-NP) and pregnant (N-P) human myometrium, as well as in human visceral (VAT) and subcutaneous (SCAT) adipose tissue (AT).MethodsFSHR expression at mRNA (real-time qPCR, RNAscope in situ hybridization) and protein (immunohistochemical staining) levels in adipose tissue, myometrium, and adipocytes were evaluated. Myometrium and adipocytes were treated with recombinant (rh)FSH to study its effects on functional pathways. Myometrium contractile activity was measured using a force transducer with digital output and the DASYLab software unit. Cyclic adenosine monophosphate (cAMP) production by myometrium explants and adipocytes was measured using a cAMP ELISA Kit. The activation of the AKT pathway in myometrium and adipocytes was analyzed by Western blot analysis.ResultsContrary to previous observations, we found no expression of FSHR at either mRNA or protein levels in M-NP, N-P, VAT, and SCAT. Treatment with recombinant human FSH (rhFSH) showed no effect on cAMP production or phosphorylation of AKT in M-NP, N-P, as well as in VAT and SCAT. rhFSH treatment did not influence contractile activity in M-NP, N-P.ConclusionsThese findings suggest that the FSHR signaling pathway does not regulate myometrial contractility during pregnancy. Additionally, the absence of FSHR expression in both VAT and SCAT implied that FSHR does not play a role in the functional signaling pathways in adipose tissues. In conclusion, our findings contradict earlier data on the involvement of FSH-FSHR signaling in regulating myometrial contractility near term, as well as in adipose tissue function.

Long Non-Coding RNA LINC01116 Promotes the Proliferation of Lung Adenocarcinoma by Targeting miR-9-5p/CCNE1 Axis.

Long non-coding RNA (lncRNA) LINC01116 is crucial in promoting cell proliferation, invasion and migration in solid tumours, including lung adenocarcinoma (LUAD). LINC01116 acts as a competing endogenous RNAs (ceRNA) that binds competitively to microRNAs and plays a critical role in tumour migration and invasion. However, other mechanisms of action besides the ceRNA theory have been rarely reported and remain to be elucidated further. The differences in RNA and protein levels in cells and tissues were assessed through real-time quantitative PCR and Western blot analysis. Invitro functional assays and invivo xenograft models were used to analyse the function of LINC01116 in LUAD. Thus, the molecular correlation between miR-9-5p and CCNE1 was investigated through direct and indirect mechanism experiments. LINC01116, miR-9-5p and CCNE1 were upregulated in LUAD cell lines and tissues and were associated with a poor prognosis in patients. LINC01116 depletion inhibited proliferation but facilitated cell apoptosis. AGO2-RNA binding protein immunoprecipitation (AGO2-RIP) experiments confirmed that AGO2 binds to LINC01116 and miR-9-5p, indicating that LINC01116 interacts with miR-9-5p. The overexpression of miR-9-5p and CCNE1 effectively counteracts the biological effects of LINC01116 knockdown on reduced proliferation and cell cycle arrest in LUAD cells. The downregulation of miR-9-5p significantly reduces the CCNE1 level in A549 cells, and the upregulation of LINC01116 counteracts the downregulation of miR-9-5p effect, restoring the expression level of CCNE1. Our data demonstrated that LINC01116 regulates the expression of CCNE1 by positively regulating miR-9-5p, thereby affecting cell cycle, proliferation and participating in the development of LUAD.

Therapeutic efficacy of CRISPR RNA nanoparticles on cervical cancer disease using ultrasound elastography imaging

This work aimed to evaluate the therapeutic efficacy of clustered regularly interspaced short palindromic repeats (CRISPR) ribonucleic acid (RNA) nanoparticles (NPs) in the therapy of cervical cancer (CC) using ultrasound elastography imaging. Using poly(β-amino esters) (PBAE), CRISPR RNA NPs encapsulating Human Papillomavirus 16 (HPV16) E7, namely Poly(β-amino ester) (PBAE)/CRISPR16E7-GFP and PBAE/shRNA-16E7, were prepared. The characterization of these NPs was conducted using transmission electron microscopy (TEM) and laser diffraction particle size analysis. The cytotoxicity of PBAE/CRISPR16E7-GFP and PBAE/shRNA-16E7 NPs on CC cell lines SiHa, HeLa, and CaSki was assessed using the cell counting kit-8 (CCK-8) assay. Sixty female Sprague Dawley (SD) rats were rolled into six groups randomlyas follows: the blank Ctrl group (Group A, no treatment), the model group (Group B, CC model), the pSpCas9-GFP plasmid group (Group C, CC model + pSpCas9-GFP), the PBAE/CRISPR-16E7 group (Group D, CC model + PBAE/CRISPR-16E7), the pSIREN-U6-shRNA-CMV-iRFP plasmid group (Group E, CC model + pSIREN-U6-shRNA-CMV-iRFP), and the PBAE/shRNA-16E7 group (Group F, CC model + PBAE/shRNA-16E7), each consisting of 10 rats. The impact of NPs on the expression levels (ELs) of HPV16E7 and RB1 proteins in tumor tissues was assessed using Western Blot (WB) analysis. Additionally, ultrasound elastography imaging was employed to analyze the strain ratio (SR) and shear wave velocity (SWV) in lesions of various rat groups. The results indicated that the PBAE/CRISPR16E7-GFP NPs exhibited a particle size of 125.64 ± 7.8 nm, a zeta potential of 19.17 ± 3.61 mV, and a polydispersity index (PDI) of 0.281 ± 0.03. These NPs demonstrated a regular spherical structure with a compact morphology. The PBAE/shRNA-16E7 NPs displayed a particle size of 112.93 ± 9.1 nm, a zeta potential of 13.54 ± 1.39 mV, and a PDI of 0.185 ± 0.03. They exhibited a uniform distribution of spherical shape with regularity in size. The plasmids within PBAE/CRISPR16E7-GFP and PBAE/shRNA-16E7 NPs exhibited distinct trends of burst and sustained release. The viability of SiHa, HeLa, and CaSki cells remained unaffected by PBAE/CRISPR16E7-GFP NPs. At various mass ratios, neglectable differences (P > 0.05) were observed in cell viability of SiHa, HeLa, and CaSki cells treated with PBAE/CRISPR16E7-GFP and PBAE/shRNA-16E7 NPs relative to Ctrl group. Relative to Group B, both Groups C and E exhibited a drastic decrease in tumor volume, tumor mass, and HPV16E7 protein ELs (P < 0.05), whereas Groups D and F showed a notably greater reduction in tumor volume, tumor mass, and HPV16E7 protein ELs (P < 0.01). The HPV16E7 protein ELs, SR, and SWV values greatly increased (P < 0.001) in tumors of Group B rats while the EL of RB1 protein notably decreased (P < 0.001) versus Group A. Groups C and E demonstrated elevated HPV16E7 ELs, SR, and SWV values (P < 0.05), with a marked decrease in RB1 protein EL (P < 0.001). Group F exhibited a notable reduction in HPV16E7 protein levels, SR, and SWV values (P < 0.001).The CRISPRRNA NPs composed of PBAE encapsulating HPV16 E7, namely PBAE/CRISPR16E7-GFP and PBAE/shRNA-16E7 plasmids, demonstrated a significant role in CC therapy. These NPs hold potential application value in the treatment of HPV-related CC, indicating their importance in addressing this particular disease.

Deciphering the prognostic landscape of triple-negative breast cancer: A focus on immune-related hub genes and therapeutic implications.

Triple-negative breast cancer (TNBC), known for its hostile nature and limited treatment modalities, has spurred researchers to explore novel approaches for enhancing clinical outcomes. Here, the study aimed to analyze transcriptomics data to identify immune-related hub genes associated with TNBC that might serve as prognostic biomarkers. Initially, we determined genes that were differentially expressed between TNBC and normal tissues by integrating microarray and RNA sequencing data. Then, through protein-protein interaction and module analysis, we identified five putative hub genes: AURKA, CCNB1, CDCA8, GAPDH, and TOP2A. Subsequently, gene ontology and Kyoto Encyclopedia of Genes and Genomes pathway analyses revealed that the hub genes were primarily involved in the progesterone-mediated oocyte maturation signaling pathway and oocyte meiosis. Additionally, we observed that these five hub genes were significantly elevated at both protein and mRNA levels in TNBC tissues and contributed to worse survival. Furthermore, the expression of these hub genes exhibited a strong positive association with immune-invading cells such as CD8 T cells, CD4 T cells, and dendritic cells. The analysis of the regulatory network revealed three transcription factors (YBX-1, E2F1, and E2F3) and three posttranscriptional regulators (hsa-mir-25-3p, hsa-mir-92a-3p, and hsa-let-7b-5p) of hub genes. Finally, we explored potential drug candidates for the hub genes using Drug-Gene Interaction Database and discovered that there are no FDA-approved drugs for CCNB1 and CDCA8, highlighting a promising area for future research. Taken together, our results will be of immense importance in addressing the intricacies of TNBC.