Cyclin D1 Research Articles

Prunella vulgaris Extract Ameliorates Testosterone-Induced Benign Prostatic Hyperplasia by Regulating Androgen Levels, Cell Proliferation, and Apoptosis

Background/Objectives: Benign prostatic hyperplasia (BPH) is a prevalent urological condition affecting elderly men. Prunella vulgaris L. (PV), a perennial herbaceous plant native to Europe and Asia, has anti-inflammatory, antioxidant, and antimicrobial effects. In this study, we determined the effect of PV extract on the development of BPH. Methods: Rats were treated via a daily hypodermic injection of testosterone propionate (TP; 3 mg/kg) for 4 weeks. Groups of BPH rats were treated with or without PV (60 or 80 mg/kg) by oral gavage. Results: In BPH model rats, PV considerably reduced their relative prostate weight and serum concentrations of dihydrotestosterone (DHT) and testosterone. The TP-induced increases in epithelial thickness in the prostate, proliferating cell nuclear antigen (PCNA) expression, and cyclin D1 expression were remarkably reduced, whereas terminal deoxynucleotidyl transferase dUTP nick-end labeling (TUNEL)-positive cells and cleaved caspase-3 levels were increased, in PV-treated rats compared to BPH rats. The mRNA expression levels of growth factors, such as transforming growth factor-β (TGF-β), fibroblast growth factor (FGF), and insulin-like growth factor (IGF-2), were significantly reduced in PV-treated rats. Mechanistically, the TP-induced activation of c-Jun N-terminal kinase (JNK) was reduced by PV administration. Conclusions: These results designate that PV effectively ameliorates the development of testosterone-induced BPH through anti-androgenic, anti-proliferative, and pro-apoptotic activities, suggesting that it could be a potential therapeutic substance for BPH.

The Impact of Acute Myeloblastic Leukemia-Derived Small Extracellular Vesicles on the Bone Marrow Mesenchymal Stem Cells: Expression of G1 Cell Cycle Progression Genes

Background: Leukemia progression is intricately linked to interactions with neighboring cells within the bone marrow microenvironment (BMM), and small extracellular vesicles (sEVs) emerge as vital mediators in facilitating these interactions. Objectives: This study examined the proliferation effects of sEVs-derived from acute myeloid leukemia (AML) in the HL60 cell line on the cell cycle progression of bone marrow mesenchymal stem cells (BM-MSCs), a key element of the BMM. Methods: Small extracellular vesicles were isolated from the HL60 cell line supernatant, using the ExoCib kit, and characterized through flow cytometry, transmission electron microscopy (TEM), dynamic light scattering (DLS), and bicinchoninic acid (BCA) protein assay. bone marrow mesenchymal stem cells were cultured and treated with various concentrations (20 μg/mL, 50 μg/mL, and 80 μg/mL) of AML-derived sEVs for 24, 48, and 72 hours. The effects on cell proliferation and viability were assessed, using methylthiazole tetrazolium test (MTT) and Ki-67 assays, while cell cycle progression and apoptosis were analyzed via flow cytometry. RT-qPCR was performed to evaluate the expression levels of CCND1, CDK4, CDK6, and AKT1 genes. Results: The proliferation effects of AML-derived sEVs on BM-MSCs were both dose- and time-dependent, with optimal effects observed at 50 μg/mL after 48 hours. Flow cytometry analysis revealed a significant increase in the G1 phase, showing a 1.8-fold change compared to the control group (P < 0.0001). RT-qPCR results demonstrated a significant upregulation of CCND1 (3.5-fold, P < 0.0001), CDK4 (3.2-fold, P < 0.0001), CDK6 (2.5-fold, P < 0.0001), and AKT1 (3.2-fold, P < 0.0001) expression levels, along with increased Ki-67 (2.3-fold, P < 0.0001) levels. Moreover, treatment with 50 μg/mL of AML-derived sEVs resulted in a notable reduction in BM-MSC apoptosis (0.57-fold decrease, P < 0.0001). Conclusions: Our findings revealed that AML extracellular vesicles could alter the gene expression associated with the proliferation of BMSCs to increase their proliferation by stimulating the cell cycle of MSCs through the G1 phase.

Repurposing of sericin combined with dactolisib or vitamin D to combat non-small lung cancer cells through computational and biological investigations

This study aims to repurpose sericin in combating non-small lung cancer cells (A549 and H460) by combining it with dactolisib or vitamin D to reduce the dose of dactolisib and boost the anticancer effectiveness of dactolisib and vitamin D. Therefore, the binding affinities of individual and combined drugs were examined using in silico and protein-protein interaction studies, targeting NF-κB, Cyclin D1, p-AKT, and VEGF1 proteins. The findings manifested remarkable affinities for combinatorial drugs compared to individual compounds. To substantiate these findings, the combined IC50 for each combination (sericin + dactolisib and sericin + vitamin D) were determined, reporting 31.9 and 41.8 µg/ml, respectively, against A549 cells and 47.9 and 55.3 µg/ml, respectively, against H460 cells. Furthermore, combination indices were assessed to lower the doses of each drug. Interestingly, in vitro results exhibited marked diminutions in NF-κB, Cyclin D1, p-AKT, and VEGF1 after treatment with sericin + dactolisib and sericin + vitamin D compared to control lung cancer cells and those treated with a single drug. Moreover, A549 and H460 cells treated with both combinations demonstrated augmented caspase-3 levels, implying substantial apoptotic activity. Altogether, these results accentuated the prospective implementation of sericin in combination with dactolisib and vitamin D at low doses to preclude lung cancer cell proliferation.

Spectrum of Gastric Carcinoma in a Tertiary Care Centre with Special Reference to PDL1 and Cyciln D1

Introduction: Gastric carcinoma is the fifth most frequently diagnosed tumor and the fourth leading cause of death. They represent a heterogenous group of tumors with multifactorial etiologies, both environmental and genetic. PD-L1 is a transmembrane protein, and a positive PD-L1 expression in gastric carcinoma may be a predictive biomarker of the response to PD-1/PD-L1 immune checkpoint inhibitors. Cyclin D1 is an important cell cycle regulatory protein and involved in the carcinogenesis of various cancers including gastric cancer. Aims: The aim of this study is to find out the clinicopathological spectrum of gastric carcinoma and to show the immunohistochemical expression of Cyclin D1 and PD-L1 to evaluate their prognostic importance and their therapeutic impact. Study Design: It is an observational cross-sectional study conducted in the Department of Pathology in collaboration with the Department of Surgery, in a tertiary care hospital. Partial and total gastrectomies were selected from January 2021 to July 2022 in a tertiary care hospital. Clinical history was collected in the study proforma. Immunohistochemical staining for Cyclin D1 and PD-L1 was performed. Material and Methods: The clinicopathological parameters like age, gender, histological type, regional lymph node status, lymphovascular invasion, perineural invasion, and pathological stage were compared and correlated with Cyclin D1 and PD-L1 expression. Results: Positive Cyclin D1 and PDL1 expression were found in 50% and 70% cases of gastric carcinoma, respectively. Cyclin D1 expression showed statistically significant correlation with anemia, regional lymph node status, and lymphovascular invasion. PD-L1 and Cyclin D1 expression showed statistically significant association between them (p value-0.0006). Conclusion: Cyclin D1 and PDL1 are important biomarkers in gastric carcinoma. Their contribution towards prediction of progression and prognosis, along with newer therapeutic modalities, is of immense importace in these patients. A statistically significant correlation was found between these two markers.

FGF6 inhibits oral squamous cell carcinoma progression by regulating PI3K/AKT and MAPK pathways

To explore diagnostic and prognostic biomarkers in the progression of oral squamous cell carcinoma (OSCC) and to reveal their regulatory mechanisms in key pathways. A RayBiotech protein chip was used to screen differentially expressed serum proteins in OSCC, oral leukoplakia (OLK), and healthy participants. Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) enrichment analysis were used to determine the pathways enriched by characteristic differential proteins. Immunohistochemical analysis and western blotting were used to verify the expression of characteristic differential proteins and key regulatory factors in human tissues and in a nude mouse model. Fibroblast growth factor 6 (FGF6) was identified as a key differential protein and was weakly expressed in OSCC tissues. The mitogen-activated protein kinases (MAPK) and PI3K-AKT pathways were identified as key signaling pathways. The results showed that pERK, Cyclin D1, pAKT, and BCL2 were highly expressed in OSCC, Caspase9 was lowly expressed in OSCC. With an increase in FGF6 expression in nude mice, the expression of FGFR4, pERK, Cyclin D1, pAKT, BCL2, GPX4, and ACSL4 increased, and the expression of Caspase9 decreased. FGF6 may change the expression of apoptosis-related proteins and proliferation factors by binding to FGFR4 in the PI3K-AKT/MAPK pathway and may inhibit the ferroptosis of OSCC, thereby possibly participating in the process of inhibiting OSCC.

ZNF652 exerts a tumor suppressor role in lung cancer by transcriptionally downregulating cyclin D3.

Dysfunction of zinc finger protein 652 (ZNF652) is associated with various malignant tumors. However, the role of ZNF652 in lung cancer (LC) is poorly understood. Here, we identified that ZNF652 was downregulated in human LC tissues and cell lines. Low ZNF652 expression was associated with poor survival in LC patients. Overexpression of ZNF652 inhibited cell viability, proliferation, migration, and invasion of LC cells, whereas ZNF652 knockdown promoted these malignant phenotypes. Using RNA-seq analysis revealed that ZNF652 overexpression resulted in obvious alterations of various biological processes, especially cell cycle and cellular senescence. Subsequently, we confirmed that ZNF652 overexpression arrested the cell cycle at the G1 phase, increased ROS-mediated DNA damage, induced LC cell senescence, and enhanced cisplatin-induced apoptosis in LC cells. Mechanistically, ZNF652 directly bound to the promoter of cyclin D3 (CCND3), inhibited its transcription, thereby arresting the cell cycle at the G1 phase. Ectopic expression of cyclin D3 rescued the decreased cell viability and cell cycle arrest induced by ZNF652. In vivo studies further showed that ZNF652 overexpression suppressed the tumorigenic potential of LC. Collectively, our findings reveal that ZNF652 exerts a tumor suppressor role in lung cancer by inducing cell cycle arrest and cellular senescence via transcriptionally downregulating cyclin D3. Thus, ZNF652 may be a prognostic predictive factor for LC patients.

TCF4 promotes apoptosis and Wnt/β-catenin signaling pathway in acute kidney injury via transcriptional regulation of COX7A2L.

Acute kidney injury (AKI) is still a serious kidney illness with high morbidity and death rates, and it's crucial to comprehend the underlying molecular causes. Bioinformatics analysis was performed on GSE139061 and GSE30718 data sets, and COX7A2L was screened out. The role of COX7A2L in H/R-treated cells and its transcriptional regulation with TCF4 were assessed. In vitro experiments analyzed the regulation of COX7A2L and TCF4 on the proliferation, apoptosis, and Wnt/β-catenin signaling pathway of H/R-treated cells. COX7A2L as a hub gene was downregulated in AKI samples. In H/R-treated cells, COX7A2L overexpression inhibited apoptosis and promoted cell proliferation, while COX7A2L knockdown promoted apoptosis and inhibited cell proliferation. Notably, TCF4 exhibited a significant positive correlation with COX7A2L. TCF4 overexpression-induced apoptosis was lessened and improved cell proliferation was countered by COX7A2L knockdown, according to rescue study findings. Besides, we discovered that TCF4 overexpression increased the expression of proteins linked to the Wnt/β-catenin signaling pathway (c-myc, β-catenin, and cyclin D1), while underexpression of COX7A2L counteracted this effect. The study revealed the pivotal role of COX7A2L in AKI, which is regulated by TCF4 and modulates the Wnt/β-catenin signaling pathway, highlighting its potential as a therapeutic target.

Versatile properties of Opuntia ficus-indica (L.) Mill. flowers: In vitro exploration of antioxidant, antimicrobial, and anticancer activities, network pharmacology analysis, and In-silico molecular docking simulation.

Opuntia ficus-indica (L.) Mill. has been used in folk medicine against several diseases. The objectives of the present study were to investigate the chemical composition of the methanolic extract of O. ficus-indica (L.) Mill. flowers and their antioxidant, antimicrobial, and anticancer properties. Besides, network pharmacology and molecular docking were used to explore the potential antitumor effect of active metabolites of O. ficus-indica (L.) Mill. against breast and liver cancer. The results revealed many bioactive components known for their antimicrobial and anticancer properties. Furthermore, scavenging activity was obtained, which indicated strong antioxidant properties. The plant extract exhibited antimicrobial activities against Aspergillus brasiliensis (MIC of 0.625 mg/mL), Candida albicans, Saccharomyces cerevisiae, Staphylococcus aureus, Escherichia coli, and Pseudomonas aeruginosa at MICs of 1.25 mg/mL. The results revealed proapoptotic activities of the O. ficus-indica (L.) Mill. extract against MCF7, MDA-MB-231, and HepG2 cell lines, where it induced significant early apoptosis and cell cycle arrest at sub-G1 phases, besides increasing the expression levels of p53, cyclin D1, and caspase 3 (p <0.005). The network pharmacology and molecular docking analysis revealed that the anticancer components of O. ficus-indica (L.) Mill. flower extract targets the PI3K-Akt pathway. More investigations might be required to test the mechanistic pathways by which O. ficus-indica (L.) Mill. might exhibit its biological activities in vivo.

Controlled release of hydrogel-encapsulated mesenchymal stem cells-conditioned medium promotes functional liver regeneration after hepatectomy in metabolic dysfunction-associated steatotic liver disease

BackgroundGlobally, prevalence of metabolic dysfunction-associated steatotic liver disease (MASLD) is increasing, and there is an urgent need to develop innovative therapies that promote liver regeneration following hepatectomy for this disease. Surgical excision is a key therapeutic approach with curative potential for liver tumors. However, hepatic steatosis can lead to delayed liver regeneration and higher post-operative complication risk. Mesenchymal stem cells-conditioned medium (MSC-CM) is considered a rich source of paracrine factors that can repair tissues and restore function of damaged organs. Meanwhile, hydrogels have been widely recognized to load MSC secretome and achieve sustained release. This study aimed to evaluate the therapeutic effect of hydrogel-encapsulated MSC-CM on liver regeneration following partial hepatectomy (PHx) in a rodent model of diet-induced hepatic steatosis.MethodsMale Lewis rats were fed with a methionine and choline–deficient diet. After 3 weeks of feeding, PHx was performed and rats were randomly allocated into two groups that received hydrogel-encapsulated MSC-CM or vehicle via the intra-mesenteric space of the superior mesenteric vein (SMV).ResultsThe regeneration of the remnant liver at 30 and 168 h after PHx was significantly accelerated, and the expressions of proliferating cell nuclear antigen were significantly enhanced in the MSC-CM group. MSC-CM treatment significantly increased hepatic ATP and β-hydroxybutyrate content at 168 h after PHx, indicating that MSC-CM fosters regeneration not only in volume but also in functionality. The number of each TUNEL- and cleaved caspase-3 positive nuclei in hepatocytes at 9 h after PHx were significantly decreased in the MSC-CM group, suggesting that MSC-CM suppressed apoptosis. MSC-CM increased serum immunoregulatory cytokine interleukin-10 and interleukin-13 at 30 h after PHx. Additionally, mitotic figures and cyclin D1 expression decreased and hepatocyte size increased in the MSC-CM group, implying that this mode of regeneration was mainly through cell hypertrophy rather than cell division.ConclusionsMSC-CM represents a novel therapeutic approach for patients with MASLD requiring PHx.

A Necessary Role for Cyclin D2 Induction During Colon Cancer Progression Mediated by L1.

The cell adhesion molecule L1CAM (L1), mainly known for its function in brain cells, is a Wnt/β-catenin signaling target gene in colorectal cancer (CRC) cells, where it promotes invasion and liver metastasis. We interrogated which genes are expressed at increased levels in human CRC tissue and induced in CRC cell lines overexpressing L1. We found increased cyclin D2 levels in CRC tissue and LS 174T and HCT 116 human CRC cells overexpressing L1. Increased cyclin D2 in CRC cells was associated with higher proliferation rates, faster motility, tumorigenesis, and liver metastasis. The suppression of cyclin D2 expression by shRNA to cyclin D2 blocked the increase in these cellular properties of L1-expressing cells. The overexpression of cyclin D2 in the absence of L1 also conferred tumorigenic properties similar to L1 expression. The pathways involved in the elevation of cyclin D2 by L1 include NF-κB, Akt, and β-catenin signaling but not the Erk pathway. We found that in a significant percentage of human CRC tissue samples, cyclin D2 is expressed at high levels in the nuclei of cancer cells. At the same time, the adjacent normal mucosa was negative for cyclin D2 staining. The results suggest that the increased cyclin D2 expression by L1 is required to induce proliferative, motile tumor development in CRC tissue and can serve as a diagnostic marker and a target for CRC therapy.

GdX inhibits the occurrence and progression of breast cancer by negatively modulating the activity of STAT3

ABSTRACT Aim To elucidate the biological functionality and regulatory mechanisms of GdX in breast cancer (BC). Methods The examination of GdX expression in human BC tissues and cell lines was conducted through immunohistochemical (IHC) and Western blot. Cell proliferation capacity was assessed via the CCK-8 and colony formation assay, while cell migration was determined through the wound healing assay. The expression levels of BCL-XL, Cyclin D1, and C-myc gene were quantified using RT-qPCR and Western blot. In vivo tumor growth was evaluated in nude mice xenografted with MDA-MB-231 cells overexpressing GdX, and a mouse model with GdX-deficient BC was established to observe the impact of GdX on BC formation and metastasis. Dual-luciferase reporter assay and immunofluorescence were employed to confirm the interaction between GdX and STAT3. Western blot was employed to validate the influence of GdX overexpression on the phosphorylation process of STAT3. Results GdX exhibited low expression in the cancer tissues of BC patients and cell lines. MDA-MB-231 and MCF-7 cells overexpressing GdX displayed a notable reduction in proliferation and diminished migratory capabilities, accompanied by downregulated mRNA and protein expression of BCL-XL, Cyclin D1, and C-myc. In the xenograft mouse model, heightened GdX expression correlated with a decelerated in vivo tumor growth. Furthermore, in mice deteleing GdX, both the quantity and weight of tumors increased, along with evident pulmonary metastasis. Mechanistically, STAT3 emerged as a downstream target gene of GdX. Conclusions GdX exerts its inhibitory effects on the initiation and progression of BC by negatively modulating the phosphorylation of STAT3.

Impaired cyclin D3 protein degradation contributes to trastuzumab resistance in HER2 positive breast cancer

As the first anti-HER2 targeted agent approved by FDA in 1998, Trastuzumab has significantly improved the outcome of patients with HER2 positive metastatic breast cancer. Unfortunately, resistance to trastuzumab is a severe obstacle to its therapeutic efficacy in clinical application, and its mechanism has not yet been fully elucidated. In our study, we found that stabilization of cyclin D3 could be one reason for trastuzumab resistance. Trastuzumab could induce G1/G0 phase arrest by downregulating cyclin D3 protein expression. However, the protein expression of cyclin D3 was not affected in trastuzumab-resistant cells, which might be related to aberrant activation of ERK signaling pathway. Furthermore, degradation of cyclin D3 protein by trastuzumab was mainly resulted from ubiquitin-dependent proteasome mechanism instead of transcriptional regulation. In trastuzumab-resistant breast cancer cells, trastuzumab-induced degradation of cyclin D3 protein was abrogated. When the ubiquitin pathway was inhibited, cells would show a predisposition to resistance to trastuzumab. Further, CDK4/6 inhibitor can inhibit the proliferation of trastuzumab-resistant HER-2 positive breast cancer cells. Therefore, combination of CDK4/6 inhibitors and anti-HER2 targeted therapy may be an alternative and promising strategy to overcome trastuzumab resistance in the future.

Evaluation of Cyclin D1 protein and its association with the clinicopathological characteristics and prognosis of lung cancer: A retrospective study from Southern Kerala, India.

Cyclin D1 is a protein that can enhance the proliferation of cancer cells and has been detected in various malignancies, including lung cancer. However, routine examinations for Cyclin D1 in lung cancer cases have not been conducted in Kerala. This study sought to evaluate the links between cyclin D1 expression, clinicopathological characteristics, and 2-year survival rates in lung cancer. This retrospective cohort study used medical records and paraffin blocks of lung cancer patients at the Regional Cancer Centre in Kerala, India, between 2015 and 2018. The data were collected from 61 subjects, comprising of lung adenocarcinoma (18%), lung squamous cell carcinoma (27.9%), non-small-cell lung carcinoma (18%), poorly differentiated carcinoma (19.7%), and negative for malignant cells (16.4%). Data analysis was conducted using SPSS. The study revealed that 31.10% of the lung cancer patients exhibited overexpression of cyclin D1. A significant correlation was observed between cyclin D1 expression and histopathological results (P = 0.002), indicating that the level of cyclin D1 might be linked to specific histopathological subtypes of lung cancer. Despite this significant finding, cyclin D1 expression did not show any association with the clinical stage of the cancer or other clinical characteristics of the patients. Furthermore, when examining the 2-year survival rates of the patients, the study found no significant difference between those who had overexpression of cyclin D1 and those who did not (P = 0.145). Cyclin D1 expression was associated with histology type of lung cancer with no significant association to prognosis.

Neuronatin Promotes the Progression of Non-small Cell Lung Cancer by Activating the NF-κB Signaling.

Understanding the regulatory mechanisms involving neuronatin (NNAT) in non-small cell lung cancer (NSCLC) is an ongoing challenge. This study aimed to elucidate the impact of NNAT knockdown on NSCLC by employing both in vitro and in vivo approaches. To investigate the role of NNAT, its expression was silenced in NSCLC cell lines A549 and H226. Subsequently, various parameters, including cell proliferation, invasion, migration, and apoptosis, were assessed. Additionally, cell-derived xenograft models were established to evaluate the effect of NNAT knockdown on tumor growth. The expression of key molecules, including cyclin D1, B-cell leukemia/lymphoma 2 (Bcl-2), p65, matrix metalloproteinase (MMP) 2, and nerve growth factor (NGF) were examined both in vitro and in vivo. Nerve fiber density within tumor tissues was analyzed using silver staining. Upon NNAT knockdown, a remarkable reduction in NSCLC cell proliferation, invasion, and migration was observed, accompanied by elevated levels of apoptosis. Furthermore, the expression of cyclin D1, Bcl-2, MMP2, and phosphorylated p65 (p-p65) showed significant downregulation. In vivo, NNAT knockdown led to substantial inhibition of tumor growth and a concurrent decrease in cyclinD1, Bcl-2, MMP2, and p-p65 expression within tumor tissues. Importantly, NNAT knockdown also led to a decrease in nerve fiber density and downregulation of NGF expression within the xenograft tumor tissues. Collectively, these findings suggest that neuronatin plays a pivotal role in driving NSCLC progression, potentially through the activation of the nuclear factor-kappa B signaling cascade. Additionally, neuronatin may contribute to the modulation of tumor microenvironment innervation in NSCLC. Targeting neuronatin inhibition emerges as a promising strategy for potential anti-NSCLC therapeutic intervention.

Female sex hormones inversely regulate acute kidney disease susceptibility throughout life.

While epidemiological and experimental studies have demonstrated kidney-protective effects of estrogen and female sex in adulthood, some epidemiological data showed deterioration of kidney function during puberty when estrogen production increases. However, molecular mechanisms explaining these conflicting phenomena remain unknown. Here, we showed that the pubertal sex hormone surge in female mice increases susceptibility to kidney ischemia reperfusion injury partly via downregulation of insulin-like growth factor 1 receptor (IGF-1R) expression in proximal tubules. Adult mice ovariectomized pre-pubertally (at postnatal day 21) showed strong tolerance to kidney ischemia, which was partly reversed by the administration of 17β-estradiol, while adult mice ovariectomized post-pubertally (at 8 weeks of age) were vulnerable to kidney ischemia. Kidney tubular IGF-1R protein expression decreased during postnatal growth but was highly expressed in adult mice ovariectomized pre-pubertally and in infant mice, which might be partly explained by different expression of an E3 ligase (MDM2) of IGF-1R. Mice deficient of Igf-1r in proximal tubules (iIGF-1RKO mice) during postnatal kidney growth showed increased susceptibility to ischemic injury. RNA-seq and western blotting analysis using proximal tubular cells from pre-pubertally ovariectomized iIGF-1RKO and control mice revealed altered expression of cell cycle-associated molecules such as cyclin D1. These results suggest that Igf-1r deletion during postnatal growth renders proximal tubular cells susceptible to ischemia possibly via altered cell cycle regulation. Thus, our findings provide evidence that exposure to pubertal sex hormones leads to increased susceptibility to kidney ischemia, which is partly mediated by modulation of IGF-1R signaling.

Alendronate sodium induces G1 phase arrest and apoptosis in human umbilical vein endothelial cells by inhibiting ROS-mediated ERK1/2 signaling

Bisphosphonates are potent bone resorption inhibitors, among which alendronate sodium (ALN) is commonly prescribed for most osteoporosis patients, but long-term application of ALN can cause bisphosphonate-related osteonecrosis of jaw (BRONJ), the pathogenesis of which remains unclear. Previous studies have suggested that bisphosphonates cause jaw ischemia by affecting the biological behavior of vascular endothelial cells, leading to BRONJ. However, the impacts of ALN on vascular endothelial cells and its mechanism remain unclear. The purpose of this work is to assess the influence of ALN on human umbilical vein endothelial cells (HUVECs) and clarify the molecular pathways involved. We found that high concentration of ALN induced G1 phase arrest in HUVECs, demonstrated by downregulation of Cyclin D1 and Cyclin D3. Moreover, high concentration of ALN treatment showed pro-apoptotic effect on HUVECs, demonstrated by increased levels of the cleaved caspase-3, the cleaved PARP and Bax, along with decreased levels of anti-apoptotic protein Bcl-2. Further experiments showed that ERK1/2 phosphorylation was decreased. Additionally, ALN provoked the build-up of reactive oxygen species (ROS) in HUVECs, leading to ERK1/2 pathway suppression. N-acetyl-L-cysteine (NAC), a ROS scavenger, efficiently promoted the ERK1/2 phosphorylation and mitigated the G1 phase arrest and apoptosis triggered by ALN in HUVECs. PD0325901, an inhibitor of ERK1/2 that diminishes the ERK1/2 phosphorylation enhanced the ALN-induced G1 phase arrest and apoptosis in HUVECs. These findings show that ALN induces G1 phase arrest and apoptosis through ROS-mediated ERK1/2 pathway inhibition in HUVECs, providing novel insights into the pathogenic process, prevention and treatment of BRONJ in individuals receiving extended use of ALN.

Association of Genetic Variants at the CDKN1B and CCND2 Loci Encoding p27Kip1 and Cyclin D2 Cell Cycle Regulators with Susceptibility and Clinical Course of Chronic Lymphocytic Leukemia

Beyond the essential role of p27Kip1 and cyclin D2 in cell cycle progression, they are also shown to confer an anti-apoptotic function in peripheral blood (PB) lymphocytes. Although the aberrant longevity and expression of p27Kip1 and cyclin D2 in leukemic cells is well documented, the exact mechanisms responsible for this phenomenon have yet to be elucidated. This study was undertaken to determine the associations between polymorphisms in the CDKN1B and CCND2 genes (encoding p27Kip1 and cyclin D2, respectively) and susceptibility to chronic lymphocytic leukemia (CLL), as well as their influence on the expression of both cell cycle regulators in PB leukemic B cells and non-malignant T cells from untreated CLL patients divided according to the genetic determinants studied. Three CDKN1B single-nucleotide polymorphisms (SNPs), rs36228499, rs34330, and rs2066827, and three CCND2 SNPs, rs3217933, rs3217901, and rs3217810, were genotyped using a real-time PCR system. The expression of p27Kip1 and cyclin D2 proteins in both leukemic B cells and non-malignant T cells was determined using flow cytometry. We found that the rs36228499A and rs34330T alleles in CDKN1B and the rs3217810T allele in the CCND2 gene were more frequent in patients and were associated with increased CLL risk. Moreover, we observed that patients possessing the CCND2rs3217901G allele had lower susceptibility to CLL (most pronounced in the AG genotype). We also noticed that the presence of the CDKN1Brs36228499CC, CDKN1Brs34330CC, CDKN1Brs2066827TT, and CCND2rs3217901AG genotypes shortened the time to CLL progression. Statistically significant functional relationships were limited to T cells and assigned to CDKN1B polymorphic variants; carriers of the polymorphisms rs34330CC and rs36228499CC (determining the aggressive course of CLL) expressed a decrease in p27Kip1 and cyclin D2 levels, respectively. We indicate for the first time that genetic variants at the CDKN1B and CCND2 loci may be considered as a potentially low-penetrating risk factor for CLL and determining the clinical outcome.

Differential expression of ORAI channels and STIM proteins in renal cell carcinoma subtypes: implications for metastasis and therapeutic targeting.

Renal cell carcinoma (RCC) presents significant clinical challenges, highlighting the importance of understanding its molecular mechanisms. While store-operated Ca2+ entry (SOCE) is known to play an essential role in tumorigenesis and metastasis, its specific implications across various RCC subtypes remain underexplored. This study analyzed SOCE-related mRNA profiles from the KIRC and KIRP projects in The Cancer Genome Atlas (TCGA) database, focusing on differential gene expression and overall survival outcomes. Functional studies in clear cell RCC (Caki-1) and papillary RCC cell lines (pRCC, Caki-2) revealed increased expression of Orai1 and Orai3, along with STIM1, exhibited in both subtypes, with decreased STIM2 and increased Orai2 expression in pRCC. Notably, Orai3 expression had a gender-specific impact on survival, particularly in females with pRCC, where it inversely correlated with STIM2 expression. Functional assays showed Orai3 dominance in Caki-2 and Orai1 in Caki- 1. Interestingly, 2-APB inhibited SOCE in Caki-1 but enhanced it in Caki-2, suggesting Orai3 as the primary SOCE channel in pRCC. Knockdown of Orai1 and Orai3 reduced cell migration and proliferation via regulating focal adhesion kinase (FAK) and Cyclin D1 in both cell lines. These findings highlight the critical roles of Orai1 and Orai3 in RCC metastasis, with Orai3 linked to poorer prognosis in females with pRCC. This study offers valuable insights into RCC diagnostics and potential therapeutic strategies targeting ORAI channels and STIM proteins.

Electric Stimulation at 448 kHz Modulates Proliferation and Differentiation of Follicle Dermal Papilla Cells

Dermal papilla cells (DPCs) regulate the hair cycle and play important roles in hair growth and regeneration. Alopecia is a pathology caused by a deregulation in the hair cycle phases. Currently, the use of physical therapies such as radiofrequency (RF) as an alternative to pharmacological treatment is increasing. Electrical stimulation by capacitive resistive electrical transfer (CRET) is one of these therapies. The objective of the present study was to analyze the effect of RF-CRET currents on DPCs. Cells were treated with subthermal 448 kHz CRET currents with two different types of signals: standard (CRET-STD) or modulated (CRET-MOD). Viability (XTT Assay), proliferation (Ki67 and ERK1/2), apoptosis (p53 and caspase 3), differentiation (β-catenin and α-SMA), and anagen markers (versican and PPARγ) were analyzed by immunofluorescence and immunoblot. CRET caused effects on the proliferation and survival of DPCs associated with increases in the expression of p-MAPK-ERK1/2, cyclin D1, and decreases in the expression of p53 and caspase 3. Also, CRET caused significant transient increases in the expression of β-catenin, involved in hair growth, and in the expression of anagen phase markers such as versican and PPARγ related to hair follicle maintenance. The present study highlights the ability of treatment with CRET therapy to cause molecular alterations in DPC involved in hair regeneration.

Pharmacological USP2 targeting suppresses ovarian cancer growth by potentiating apoptosis and ferroptosis

Ovarian cancer is a frequently observed type of gynaecologic malignancy generally associated with poor prognosis around the world. Ubiquitin-specific proteases (USPs) form the largest subfamily of deubiquitylating enzymes and have emerged as potential therapeutic targets against human cancers. Through a systematic analysis of the prognostic significance of USP expression, USP2 was found to be inversely correlated with patient survival in ovarian cancer. Accordingly, we investigated the effects of pharmacological inhibition of USP2 on ovarian cancer by exploiting its small molecule inhibitor ML364. Our findings show that ML364 effectively hindered ovarian cancer growth and migration using a series of in vitro assays. In addition to apoptosis induction, ML364 also sensitized ovarian cancer cells to ferroptosis. Mechanistically, ML364 treatment resulted in cyclin D1 downregulation, increased poly (ADP-ribose) polymerase (PARP) cleavage, and elevated ROS levels in ovarian cancer cells. Collectively, our findings suggest USP2 as a potential therapeutic target in ovarian cancer, and hence, its pharmacological inhibition warrants further investigation.