Expression Levels Of Ki-67 Research Articles

Knockdown of HNRNPM inhibits the progression of glioma through inducing ferroptosis

ABSTRACT Purpose Ferroptosis acts as an important regulator in diverse human tumors, including the glioma. This study aimed to screen potential ferroptosis-related genes involved in the progression of glioma. Materials and Methods Differently expressed genes (DEGs) were screened based on GSE31262 and GSE12657 datasets, and ferroptosis-related genes were separated. Among the important hub genes in the protein-protein interaction networks, HNRNPM was selected as a research target. Following the knockdown of HNRNPM, the viability, migration, and invasion were detected by CCK8, wound healing, and transwell assays, respectively. The role of HNRNPM knockdown was also verified in a xenograft tumor model in mice. Immunohistochemistry detected the expression levels of HNRNPM and Ki67. Moreover, the ferroptosis was evaluated according to the levels of iron, glutathione peroxidase (GSH), and malondialdehyde (MDA), as well as the expression of PTGS2, GPX4, and FTH1. Results Total 41 overlapping DEGs relating with ferroptosis and glioma were screened, among which 4 up-regulated hub genes (HNRNPM, HNRNPA3, RUVBL1, and SNRPPF) were determined. The up-regulation of HNRNPM presented a certain predictive value for glioma. In addition, knockdown of HNRNPM inhibited the viability, migration, and invasion of glioma cells in vitro, and also the tumor growth in mice. Notably, knockdown of HNRNPM enhanced the ferroptosis in glioma cells. Furthermore, HNRNPM was positively associated with SMARCA4 in glioma. Conclusions Knockdown of HNRNPM inhibits the progression of glioma via inducing ferroptosis. HNRNPM is a promising molecular target for the treatment of glioma via inducing ferroptosis. We provided new insights of glioma progression and potential therapeutic guidance.

Dysregulated Expression of LncRNA-SChLAP1 in Breast Cancer

This study aimed to investigate the expression and clinical significance of the long chain non-coding RNA SCHLAP1 in breast cancer tissues. The research included 60 breast cancer patients treated between June 2017 and September 2019. Cancer and adjacent tissues were collected for analysis. Furthermore, breast cancer cell lines MCF-7 and HCC1937, along with normal breast epithelial cell line MCF10A, were used to study the impact of LncRNA SCHLAP1 on breast cancer cell phenotypes. qRT-PCR was employed to measure LncRNA SCHLAP1 expression levels in cells and tissues. The results demonstrated that LncRNA SCHLAP1 was significantly up-regulated in breast cancer cells and patient tissues (P <0.01). Moreover, differences in LncRNA SCHLAP1 expression were observed in patients with varying age, lymph node invasion, TNM staging, HER-2, and Ki-67 expression levels (P <0.01). Patients with high LncRNA SCHLAP1 expression had a significantly lower two-year survival rate (P <0.01). In vitro experiments revealed that down-regulated LncRNA SCHLAP1 inhibited the proliferation, migration, and invasion of MCF-7 cells, while promoting apoptosis (P <0.01). This study suggests that LncRNA SCHLAP1 is associated with breast cancer progression and patient survival, serving as an independent predictor for breast cancer progression.

Deep Learning to Predict the Cell Proliferation and Prognosis of Non-Small Cell Lung Cancer Based on FDG-PET/CT Images.

(1) Background: Cell proliferation (Ki-67) has important clinical value in the treatment and prognosis of non-small cell lung cancer (NSCLC). However, current detection methods for Ki-67 are invasive and can lead to incorrect results. This study aimed to explore a deep learning classification model for the prediction of Ki-67 and the prognosis of NSCLC based on FDG-PET/CT images. (2) Methods: The FDG-PET/CT scan results of 159 patients with NSCLC confirmed via pathology were analyzed retrospectively, and the prediction models for the Ki-67 expression level based on PET images, CT images and PET/CT combined images were constructed using Densenet201. Based on a Ki-67 high expression score (HES) obtained from the prediction model, the survival rate of patients with NSCLC was analyzed using Kaplan-Meier and univariate Cox regression. (3) Results: The statistical analysis showed that Ki-67 expression was significantly correlated with clinical features of NSCLC, including age, gender, differentiation state and histopathological type. After a comparison of the three models (i.e., the PET model, the CT model, and the FDG-PET/CT combined model), the combined model was found to have the greatest advantage in Ki-67 prediction in terms of AUC (0.891), accuracy (0.822), precision (0.776) and specificity (0.902). Meanwhile, our results indicated that HES was a risk factor for prognosis and could be used for the survival prediction of NSCLC patients. (4) Conclusions: The deep-learning-based FDG-PET/CT radiomics classifier provided a novel non-invasive strategy with which to evaluate the malignancy and prognosis of NSCLC.

Is Ki-67 Really Useful as a Predictor for Response to Neoadjuvant Chemotherapy in Locally Advanced Breast Cancer?

Neoadjuvant chemotherapy (NACT) is routinely offered to operable locally advanced breast cancer (LABC) patients desirous of breast conservation surgery and inoperable LABC patients. Pathological complete response (pCR) following chemotherapy is recognized as a surrogate for survival outcomes in high grade tumour subtypes. Many biological and tumor characters have been shown to predict pCR. The current study was performed with the aim of investigating the ability of Ki-67 in predicting pCR with NACT in breast cancer patients. A total of 105 patients with locally advanced breast cancer who completed NACT followed by surgery were included in this study from January 2020 till December 2022. Patients with advanced metastatic breast carcinoma, who did not give consent for NACT, who did not complete NACT and who did not undergo surgery were excluded. All patients were assessed for Ki-67 score on core-needle biopsy samples and response rate was assessed clinically and by histopathological examination of resected specimen. Quantitative variables were compared using unpaired t-test or Mann-Whitney 'U' test and for categorical variables Chi-square or Fisher's exact test were used. Receiver operating characteristic (ROC) curve analysis was performed to assess the predictive potential of Ki-67 expression levels in predicting pCR. To identify the predictive factors associated with pCR, univariate analysis was performed. The P value < 0.05 was considered as statistically significant. Mean age was 51.57 ± 10.8years. 51 patients achieved clinical complete response (cCR) and 33 achieved pCR after NACT. Mean Ki-67 index in overall study population, in pCR group and no pCR group was 46.44 ± 22.92%, 51.60 ± 22.3% and 44.06 ± 22.7%, respectively. On univariate analysis, ER negativity, PR negativity and Her 2neu positivity were found predictive of pCR. On subgroup analysis, TNBC and Her 2neu positive sub groups were associated with higher cCR and pCR rate. We found no significant association between Ki-67 and pCR. This result may be confounded by the fact that a significant duration of the study was in the COVID-19 pandemic. Validation of this data is required in a large prospective study.

Modeling of FAN1-Deficient Kidney Disease Using a Human Induced Pluripotent Stem Cell-Derived Kidney Organoid System.

Karyomegalic interstitial nephritis (KIN) is a genetic kidney disease caused by mutations in the FANCD2/FANCI-Associated Nuclease 1 (FAN1) gene on 15q13.3, which results in karyomegaly and fibrosis of kidney cells through the incomplete repair of DNA damage. The aim of this study was to explore the possibility of using a human induced pluripotent stem cell (hiPSC)-derived kidney organoid system for modeling FAN1-deficient kidney disease, also known as KIN. We generated kidney organoids using WTC-11 (wild-type) hiPSCs and FAN1-mutant hiPSCs which include KIN patient-derived hiPSCs and FAN1-edited hiPSCs (WTC-11 FAN1+/-), created using the CRISPR/Cas9 system in WTC-11-hiPSCs. Kidney organoids from each group were treated with 20 nM of mitomycin C (MMC) for 24 or 48 h, and the expression levels of Ki67 and H2A histone family member X (H2A.X) were analyzed to detect DNA damage and assess the viability of cells within the kidney organoids. Both WTC-11-hiPSCs and FAN1-mutant hiPSCs were successfully differentiated into kidney organoids without structural deformities. MMC treatment for 48 h significantly increased the expression of DNA damage markers, while cell viability in both FAN1-mutant kidney organoids was decreased. However, these findings were observed in WTC-11-kidney organoids. These results suggest that FAN1-mutant kidney organoids can recapitulate the phenotype of FAN1-deficient kidney disease.

Effect of trichostatin A on biological characteristics of side population cells of cervical cancer cell line (HeLa)

Purpose: To investigate the effect of trichostatin A (TSA) on biological characteristics of side population (SP) cells of cervical cancer cell line (HeLa)Methods: Side population (SP) and NSP cells were obtained from 6th generation of primary cervical cancer cells and cervical cancer cell line (HeLa). Cell surface markers (ATP-binding membrane transporter superfamily G member 2 (ABCG2), CD133, CD43, p63, Ki67, multidrug resistance (MDR) as well as cell cycle phase distribution and apoptosis, were determined. SP cells in HeLa were divided into 3 groups that received TSA at doses of 0.01, 0.05, and 0.2 μM. Untreated cells served as control. Cell growth inhibition rates of different dose groups were compared. SP cells in HeLa were divided into simple irradiation group and combined irradiation group TSA (10 % inhibition concentration (IC10) + irradiation, and values of SP cell survival fraction (SF) of the two groups under different irradiation conditions were compared.Results: The content of SP in HeLa cell line was significantly higher than in cervical cancer tissue (p &gt; 0.05). There were no significant differences in expression levels of ABCG2, CD133, CD43, p63, Ki67, and MDR, as well as G0/G1, S, and G2/M phase distributions and apoptosis rate between HeLa cell line and cervical cancer tissue (p &gt; 0.05). On the 3rd, 5th, and 7th day, SF value of SP cells was significantly higher in NSP cells (p &gt; 0.05). With increasing concentration of TSA, SF value of NSP cells gradually decreased, while that of SP cells did not change significantly.Conclusion: Cervical cancer cell line (HeLa) contains SP cells which have biological characteristics of tumor stem cells. Moreover, TSA exerts good cytotoxicity and radio-sensitization effect on SP cells.

Astragaloside IV Suppresses the Effects of Hepatocellular Carcinoma Cells on Proliferation, Angiogenesis, and Invasion in Human Umbilical Vein Endothelial Cells by Controlling Exosomes by Inhibiting Rab27a

Hepatocellular carcinoma (HCC) is the third most common cancer worldwide, and invasion and metastasis are the leading causes of death. Astragaloside IV (AS-IV), a kind of traditional Chinese medicine, has been widely used in cancer therapy. Cell Counting Kit-8, tube formation, and Transwell analyses were performed to determine cell proliferation, invasion, and angiogenesis of human umbilical vein endothelial cells (HUVECs) under the treatment of Hep3B or SK-Hep1 cells-derived exosomes or Hep3B or SK-Hep1 cells with or without AS-IV treatment. Then, Rab27a overexpression in Hep3B cells was selected to determine the function of AS-IV on the regulation of cell proliferation, invasion, and angiogenesis of HUVECs. Ultimately, the injection of Hep3B cell-derived exosomes or treatment with AS-IV on nude mice was used to comprehensively investigate AS-IV functions in vivo. Hep3B or SK-Hep1 cells promoted proliferation, invasion, and angiogenesis of HUVECs, which were inhibited by AS-IV therapy. The concentrations of Hep3B and SK-Hep1 cells-derived exosomes were markedly reduced after AS-IV therapy. Meanwhile, Rab27a expression was significantly decreased in SK-Hep1 or Hep3B cells after AS-IV therapy, and it inhibited the effects of AS-IV on Hep3B-induced proliferation, invasion, and angiogenesis of HUVECs. Tumor growth was also suppressed by AS-IV, which was accompanied by the decreased expression levels of Ki67 and CD34. Overall, this study demonstrated that AS-IV inhibits the effects of HCC cell-derived exosomes on proliferation, invasion, and angiogenesis of HUVECs via inhibiting Rab27a expression. Practical Applications. To our knowledge, this is the first study to explore the efficacy of AS-IV against HCC cell-induced proliferation, invasion, and angiogenesis of HUVECs to reveal the underlying mechanism. In this study, CCK-8, Transwell, and tube formation assay detection show that AS-IV has an inhibitory effect on HCC cell-induced proliferation, invasion, and angiogenesis of HUVECs by inhibiting Rab27a, which helps in the development of novel nutraceutical/functional food against HCC progression and thus could improve the quality of life in patients with HCC.

CT radiomics model for predicting the Ki-67 proliferation index of pure-solid non-small cell lung cancer: a multicenter study.

This study aimed to explore the efficacy of the computed tomography (CT) radiomics model for predicting the Ki-67 proliferation index (PI) of pure-solid non-small cell lung cancer (NSCLC). This retrospective study included pure-solid NSCLC patients from five centers. The radiomics features were extracted from thin-slice, non-enhanced CT images of the chest. The minimum redundancy maximum relevance (mRMR) and least absolute shrinkage and selection operator (LASSO) were used to reduce and select radiomics features. Logistic regression analysis was employed to build predictive models to determine Ki-67-high and Ki-67-low expression levels. Three prediction models were established: the clinical model, the radiomics model, and the nomogram model combining the radiomics signature and clinical features. The prediction efficiency of different models was evaluated using the area under the curve (AUC). A total of 211 NSCLC patients with pure-solid nodules or masses were included in the study (N=117 for the training cohort, N=49 for the internal validation cohort, and N=45 for the external validation cohort). The AUC values for the clinical models in the training, internal validation, and external validation cohorts were 0.73 (95% CI: 0.64-0.82), 0.75 (95% CI:0.62-0.89), and 0.72 (95% CI: 0.57-0.86), respectively. The radiomics models showed good predictive ability in diagnosing Ki-67 expression levels in the training cohort (AUC, 0.81 [95% CI: 0.73-0.89]), internal validation cohort (AUC, 0.81 [95% CI: 0.69-0.93]) and external validation cohort (AUC, 0.78 [95% CI: 0.64-0.91]). Compared to the clinical and radiomics models, the nomogram combining both radiomics signatures and clinical features had relatively better diagnostic performance in all three cohorts, with the AUC of 0.83 (95% CI: 0.76-0.90), 0.83 (95% CI: 0.71-0.94), and 0.81 (95% CI: 0.68-0.93), respectively. The nomogram combining the radiomics signature and clinical features may be a potential non-invasive method for predicting Ki-67 expression levels in patients with pure-solid NSCLC.

Intratumoral and peritumoral radiomics model based on abdominal ultrasound for predicting Ki-67 expression in patients with hepatocellular cancer.

Hepatocellular cancer (HCC) is one of the most common tumors worldwide, and Ki-67 is highly important in the assessment of HCC. Our study aimed to evaluate the value of ultrasound radiomics based on intratumoral and peritumoral tissues in predicting Ki-67 expression levels in patients with HCC. We conducted a retrospective analysis of ultrasonic and clinical data from 118 patients diagnosed with HCC through histopathological examination of surgical specimens in our hospital between September 2019 and January 2023. Radiomics features were extracted from ultrasound images of both intratumoral and peritumoral regions. To select the optimal features, we utilized the t-test and the least absolute shrinkage and selection operator (LASSO). We compared the area under the curve (AUC) values to determine the most effective modeling method. Subsequently, we developed four models: the intratumoral model, the peritumoral model, combined model #1, and combined model #2. Of the 118 patients, 64 were confirmed to have high Ki-67 expression while 54 were confirmed to have low Ki-67 expression. The AUC of the intratumoral model was 0.796 (0.649-0.942), and the AUC of the peritumoral model was 0.772 (0.619-0.926). Furthermore, combined model#1 yielded an AUC of 0.870 (0.751-0.989), and the AUC of combined model#2 was 0.762 (0.605-0.918). Among these models, combined model#1 showed the best performance in terms of AUC, accuracy, F1-score, and decision curve analysis (DCA). We presented an ultrasound radiomics model that utilizes both intratumoral and peritumoral tissue information to accurately predict Ki-67 expression in HCC patients. We believe that incorporating both regions in a proper manner can enhance the diagnostic performance of the prediction model. Nevertheless, it is not sufficient to include both regions in the region of interest (ROI) without careful consideration.

Quantitative parameters in novel spectral computed tomography for assessing gastric cancer and cell proliferation

ObjectivesTo investigate the diagnostic ability of novel spectral CT-derived parameters for gastric cancer histological types and Ki-67 expression. MethodsA total of 72 patients with histologically proven gastric cancer (GC) were retrospectively included in this study. All patients underwent dual-phase enhanced abdominal spectral CT. The arterial (AP) and venous phase (VP) slope of the spectral curve (λHU), iodine concentration (IC), normalized IC (NIC), effective atomic number (Zeff) and iodine-no-water concentration were retrospectively compared between patients with low and high Ki-67 expression levels and with different histological types in GC patients. The ROI was outlined independently by two senior physicians, and the average of three measurements at the largest level was taken. In addition, interobserver reproducibility was assessed by Bland-Altman analysis. Correlations between quantitative parameters and Ki-67 expression levels were assessed by Spearman's correlation coefficients. ResultsThe values between the mucinous group and nonmucinous carcinoma group were significantly different in both phases. The IC, NIC, and iodine-no-water concentration in the VP were significantly different among the Ki-67_L, Ki-67_M, and Ki-67_H groups. Spearman rank correlation analysis demonstrated a positive correlation between Ki-67 expression levels and IC, NIC, and iodine-no-water concentration in the VP, with correlation coefficients of 0.304, 0.424, and 0.322, respectively. ConclusionQuantitative spectral parameters can discriminate between low and high Ki-67 expression and different histological types in GC. The NIC, IC and iodine-no-water concentration can be useful parameters for evaluating of Ki-67 expression levels.

CS-NO suppresses inhibits glycolysis and gastric cancer progression through regulating YAP/TAZ signaling pathway.

Gastric cancer (GC) is a significant contributor to global mortality and is recognized for its elevated prevalence and fatality rates. Nitric Oxide (NO) plays a role in multiple aspects of cancer metastasis and progression. CS-NO is a polysaccharide-based biomaterial with NO-releasing properties that shows promising therapeutic potential. Nonetheless, the action mechanism of CS-NO in GC is still largely unclear. The present study employed various experimental techniques, including CCK-8 assay, colony formation assay, EdU staining, and transwell assays, to evaluate the proliferation, migration, and invasion of GC cells. Additionally, ELISA was utilized to measure glucose uptake, lactate production, and cellular ATP levels in GC cells. In vivo investigations on nude mice were conducted to validate the in vitro results. The present study aimed to examine the potential anti-tumor properties of CS-NO on GC through in vitro and in vivo investigations, while also exploring the underlying mechanisms involved. Our data suggested that CS-NO might prevent GC cell invasion and migration. Decreased expressions of GLUT1, HK2, and LDHA further demonstrated that CS-NO significantly suppressed aerobic glycolysis in GC cells. The administration of CS-NO resulted in a significant reduction of YAP and TAZ levels in GC cells. Our data further show that CS-NO treatment could inhibit GC cancer growth in mice, consistent with the significant decrease in Ki67, GLUT1 and YAP expression levels. These findings could reveal the good effects of CS-NO therapy on inhibiting GC.

IMMUNOHISTOCHEMICAL ASSESSMENT OF PROLIFERATION AND APOPTOSIS OF KERATINOCYTES AFTER A SINGLE FRACTIONAL IONIZING IRRADIATION

Radiation therapy for non-melanoma skin cancers is used by oncologists and radiologists in cases of ineffective surgical treatment and as adjuvant or palliative therapy. The creation of experimental models to study the proliferative-apoptotic balance of keratinocytes makes it possible to assess the degree and depth of post-radiation skin damage after exposure to electrons in order to select optimal doses in the clinic. The purpose of the study: immunohistochemical assessment of proliferation and apoptosis of keratinocytes after local electron irradiation of the skin. The experiment used male Wistar rats (n=50), which were divided into groups: I – control (n=20), which were injected with saline solution, and experimental rats, which received local electron irradiation of the skin of the outer surface of the thigh: II – 8 Gy (n=10; single), III – 40 Gy (n=10, single), IV – fractional 13 Gy for 6 days with a total focal dose of 78 Gy (n=10;). After completion of the experiment, a skin biopsy of the outer surface of the thigh was performed and an immunohistochemical study of histological preparations with antibodies to the Ki-67, caspase-3 and p53 antigens was performed. Analysis of skin fragments after irradiation showed a shift in the proliferative-apoptotic balance towards keratinocyte apoptosis: a decrease in the level of Ki-67 expression and an increase in the number of p53-positive cells. After a single irradiation with electrons at doses of 8 Gy and 40 Gy, a significant increase in keratinocytes stained positively with antibodies to caspase-3 was found - by 3.4 and 6.6 times, respectively, and with a fractional irradiation mode at a total dose of 78 Gy - by 11.6 times compared to the control group. Thus, local irradiation with electrons in single doses of 8 Gy and 40 Gy leads to a shift in the proliferative-apoptotic balance of keratinocytes towards their apoptosis, the activity of which is directly proportional to the dose of ionizing radiation, and the fractional mode (total focal dose of 78 Gy) leads to partial desquamation of the epithelium and inflammatory infiltration.

Ipatasertib exhibits anti‑tumorigenic effects and enhances sensitivity to paclitaxel in endometrial cancer invitro and invivo.

Endometrial cancer is the most common gynecologic cancer and one of the only cancers for which incidence and mortality is steadily increasing. Although curable with surgery in the early stages, endometrial cancer presents a significant clinical challenge in the metastatic and recurrent setting with few novel treatment strategies emerging in the past fifty years. Ipatasertib (IPAT) is an orally bioavailable pan‑AKT inhibitor, which targets all three AKT isoforms and has demonstrated anti‑tumor activity in pre‑clinical models, with clinical trials emerging for many cancer types. In the present study, the MTT assay was employed to evaluate the therapeutic efficacy of IPAT or IPAT in combination with paclitaxel (PTX) in endometrial cancer cell lines and primary cultures of endometrial cancer. The effect of IPAT and PTX on the growth of endometrial tumors was evaluated in a transgenic mouse model of endometrial cancer. Apoptosis was assessed using cleaved caspase assays and cellular stress was assessed using ROS, JC1 and tetramethylrhodamine ethyl ester assays. The protein expression levels of markers of apoptosis and cellular stress, and DNA damage were evaluated using western blotting and immunohistochemistry. IPAT significantly inhibited cell proliferation, caused cell cycle G1 phase arrest, and induced cellular stress and mitochondrial apoptosis in a dose dependent manner in human endometrial cancer cell lines. Combined treatment with low doses of IPAT and PTX led to synergistic inhibition of cell proliferation and induction of cleaved caspase 3 activity in the human endometrial cancer cell lines and the primary cultures. Furthermore, IPAT effectively reduced tumor growth, accompanied by decreased protein expression levels of Ki67 and phosphorylation of S6 in the Lkb1fl/flp53fl/fl mouse model of endometrioid endometrial cancer. The combination of IPAT and PTX resulted in increased expression of phosphorylated‑H2AX and KIF14, markers of DNA damage and microtubule dysfunction respectively, as compared with IPAT alone, PTX alone or placebo‑treated mice. The results of the present study provide a biological rationale to evaluate IPAT and the combination of IPAT and PTX in future clinical trials for endometrial cancer.

Impacts of LncRNA NORAD on the Proliferation, Apoptosis, and Chemosensitivity of Non-small Cell Lung Cancer Cells by Regulating ZNF217 through MiR-199a-3p

The treatment and diagnosis of non-small cell lung cancer (NSCLC) is still a difficult problem in the medical community, and exploring the molecular mechanism of the occurrence and development of NSCLC is a hot topic of the current research. Long non-coding RNA (lncRNA) NORAD is highly expressed in a variety of cancer cells. It may be a molecular target that promotes NSCLC. The aim of this study was to investigate the impacts of lncRNA NORAD on the proliferation, apoptosis, and chemosensitivity of NSCLC by regulating zinc finger protein 217 (ZNF217) through miR-199a-3p. Real-time quantitative polymerase chain reaction (qRT-PCR) method was applied to detect the expressions of NORAD, miR-199a-3p and ZNF217 genes in normal lung epithelial cells BEAS-2B, lung cancer H460 cells, and Cisplatin (DDP) resistant cell lines H460/DDP. H460/DDP cells were devided into control group, si-NC group, si-NORAD group, miR-NC group, miR-199a-3p mimic group, si-NORAD+inhibitor NC group and si-NORAD+miR-199a-3p inhibitor group. Cell proliferation, apoptosis, the expression of NORAD, miR-199a-3p and ZNF217 genes of cells in each group were detected and the expression levels of Ki-67, caspase-9 and ZNF217 proteins in different cells were also observed. The relationship between miR-199a-3p, NORAD and ZNF217 was vefified. Compared with BEAS-2B cells, the expressions of NORAD, ZNF217 mRNA were significantly increased in H460 and H460/DDP cells (P<0.05) but the expression of miR-199a-3p was significantly reduced (P<0.05). Compared with H460 cells, the expression of NORAD and ZNF217 mRNA in H460/DDP cells was significantly increased (P<0.05) and the expression of miR-199a-3p was significantly reduced (P<0.05). Compared with the control group and si-NC group, the proliferation rate, NORAD and ZNF217 mRNA expression, Ki-67 and ZNF217 protein expression of H460/DDP cells in the si-NORAD group were significantly reduced (P<0.05), but the apoptosis rate, miR-199a-3p expression and caspase-9 expression were significantly increased (P<0.05). Compared with the miR-NC group, the proliferation rate, NORAD and ZNF217 mRNA expression, Ki-67 and ZNF217 protein expression of H460/DDP cells in the miR-199a-3p mimic group were significantly reduced (P<0.05), but the apoptosis rate, miR-199a-3p expression and caspase-9 expression were significantly increased (P<0.05). Compared with the si-NORAD+inhibitor NC group, the proliferation rate, ZNF217 mRNA expression, Ki-67 and ZNF217 protein expression of H460/DDP cells in the si-NORAD+miR-199a-3p inhibitor group were significantly increased (P<0.05), the apoptosis rate, miR-199a-3p expression and caspase-9 expression were obviously increased reduced (P<0.05). Down-regulating NORAD expression can enhance miR-199a-3p expression and inhibit ZNF217 expression, thereby inhibiting H460/DDP cell proliferation, promoting apoptosis and enhancing its DDP chemotherapy sensitivity.

Jiawei Shoutai Pill promotes decidualization by regulating the SGK1/ENaC pathway in recurrent spontaneous abortion

Ethnopharmacological relevanceJiawei Shoutai Pill (JWSTW) is a traditional herbal formula for recurrent spontaneous abortion (RSA). Although JWSTW significantly improves the clinical symptoms of RSA patients, its molecular mechanism remains unclear. Aim of studyThis study evaluated the expression and function of the serum/glucocorticoid regulated kinase 1/epithelial sodium channel (SGK1/ENaC) pathway and decidualization level in RSA patients and mice. It also investigated the therapeutic effects and potential mechanisms of JWSTW. Materials and methods30 early RSA patients and 30 normal pregnant women undergoing induced abortion during the same period were included in the study. Decidual tissues were collected, and HE staining, immunohistochemistry, Western blot, and RT-PCR were used to detect protein and mRNA expression levels of SGK1, ENaC-a, estrogen Rreceptor β (ERβ), and progesterone receptor (PR) in patients’ decidual tissues. Protein expression levels of prolactin receptor (PRLR) and insulin-like growth factor binding protein 1 (IGFBP-1) were also detected. A classical RSA mouse model was constructed, and the mice were randomly divided into four groups: normal, model, dydrogesterone (DQYT) (0.33 g/kg/d), and JWSTW (1.66 g/kg/d). The normal and model groups received the same volume of distilled water by gavage for 8 and 14 days after pregnancy. On the 14th day of pregnancy, the embryonic loss rate of each group, the number of offspring born to naturally delivered mice, and the protein or mRNA expression levels of key factors of the SGK1/ENaC pathway (SGK1, ENaC-a, ERβ, and PR), decidual proliferation marker (Ki67), mesenchymal-epithelial transition (E-cadherin and Vimentin), and decidualization markers (PRLR and IGFBP-1) in mouse decidual tissue on the eighth day of pregnancy were observed. ResultsThe decidual tissue structure of RSA patients was abnormal. Immunohistochemical analysis revealed significantly reduced positive expression of SGK1, ENaC-a, ERβ, and PR proteins in the decidual tissue of RSA patients (P < 0.001). Western blot and RT-PCR analyses demonstrated significantly decreased protein and mRNA expression of SGK1, ENaC-a, ERβ, and PR in the decidual tissue of RSA patients (all P < 0.05). Additionally, protein expression of PRLR and IGFBP-1 was significantly reduced (both P < 0.001). The RSA mouse model exhibited a significant increase in embryo loss rate and decreased litter size (both P < 0.001). Treatment with DQYT and JWSTW rescued the embryo loss rate and litter size to varying extents (all P < 0.05). The protein or mRNA expression levels of SGK1, ENaC-a, ERβ, PR, Ki67, E-cadherin, vimentin, PRLR, and IGFBP-1 in RSA mice were improved to different degrees after treatment with DQYT and JWSTW (all P < 0.05). ConclusionsAbnormal SGK1/ENaC signaling pathway regulation is closely associated with early endometrial damage in RSA patients. JWSTW promotes endometrial proliferation and mesenchymal-epithelial transition through the SGK1/ENaC signaling pathway, improving endometrial shedding. Consequently, JWSTW is a potential treatment for RSA.

Studying Luminal, A and B subtypes of the breast cancer under paracrine secretion of fibroblasts

Introduction Understanding the role of the tumor microenvironment in specifying molecule markers of breast cancer subtypes could involve tumor microenvironment properties in subtyping and clinical assessment of breast cancer. Therefore, interconversion of luminal states and or their specific markers alteration under the control of TME particularly CAFs deserve to be more investigated. Methods To activate normal human fibroblasts, liquid overlay technique or nemesis was used and α-SMA expression, CAFs marker, in fibroblastic spheroids was measured by blotting. The luminal A, MCF-7, and luminal B, MDA-MB 361, cell lines are treated with normal and spheroidal/activated fibroblast conditioned medium for 48 hours. The morphological changes of luminal A and B cells were evaluated by invert microscopy and analyzed by the shape factor formula. Moreover, sensitivity changes to chemotherapeutic agents, Ki67 expression, and gene expression level including ER-related and proliferative genes were assessed respectively by MTT assay, Immunofluorescence, and real time qRT-PCR methods. Results Activated fibroblast, spheroid form, by LOT/nemosis expressed α-SMA marker two folds more than monolayer cultured fibroblasts. Our study underlined a significant increase in IC50 of both luminal A and B cell lines after being co-cultured with conditioned medium markedly in spheroidal conditioned medium. Morphological changes study by shape factor formula demonstrated more aggressiveness with gaining mesenchymal features both in luminal A and B subtypes by increasing exposure time. Changes in Ki67 expression level were observed after fibroblasts and spheroidal paracrine conditioning. Only this data supports the luminal interconversion by elevated Ki67 expression in luminal A and lowered Ki67 expression in luminal B. Gene expression results revealed that the MCF7 and MDA-MB cells as representatives luminal A and B subtypes in this study did not interchange their own specific gene pattern, however both luminal types in co-culture with condition medium increased the expression of anti-apoptotic Bcl2 gene. Conclusion Under the paracrine influence of fibroblast cells, both luminal A(MCF7) and B (MDA-MB) subtypes of breast cancer gained invasive, anti-apoptotic, and chemoresistance features which are mostly increased by activated fibroblast conditioned medium mimicking CAFs. There was no strong proof for interconversion of luminal A and B, with more similarities, subtypes of breast cancer.

Automatic analysis framework based on 3D-CT multi-scale features for accurate prediction of Ki67 expression levels in substantial renal cell carcinoma

PurposeTo investigate the effectiveness of an automatic analysis framework based on 3D-CT multi-scale features in predicting Ki67 expression levels in substantial renal cell carcinoma (RCC).MethodsThis retrospective study was conducted using multi-center cohorts consisting of 588 participants with pathologically confirmed RCC. The participants were divided into an internal training set (n = 485) and an external testing set (n = 103) from four and one local hospitals, respectively. The proposed automatic analytic framework comprised a 3D kidney and tumor segmentation model constructed by 3D UNet, a 3D-CT multi-scale features extractor based on the renal–tumor feature, and a low or high Ki67 prediction classifier using XGBoost. The framework was validated using a fivefold cross-validation strategy. The Shapley additive explanation (SHAP) method was used to determine the contribution of each feature.ResultsIn the prediction of low or high Ki67, the combination of renal and tumor features achieved better performance than any single features. Internal validation using a fivefold cross-validation strategy yielded AUROC values of 0.75 ± 0.1, 0.75 ± 0.1, 0.83 ± 0.1, 0.77 ± 0.1, and 0.87 ± 0.1, respectively. The optimal model achieved an AUROC of 0.87 ± 0.1 and 0.82 ± 0.1 for low vs. high Ki67 prediction in the internal validation and external testing sets, respectively. Notably, the tumor first-order-10P was identified as the most influential feature in the model decision.ConclusionsOur study suggests that the proposed automatic analysis framework based on 3D-CT multi-scale features has great potential for accurately predicting Ki67 expression levels in substantial RCC.Critical relevance statementAutomatic analysis framework based on 3D-CT multi-scale features provides reliable predictions for Ki67 expression levels in substantial RCC, indicating the potential usage of clinical applications.Graphical abstract

Association of the Estrogen Receptors Beta Expression with the Ki-67 Proliferative Index in Breast Cancer

Background. Estrogen receptors beta (ERβ) are an important biological regulator and target of antiestrogens, however, unlike estrogen receptors alpha (ERɑ), their significance in the prognosis and treatment of breast cancer remains unclear. Purpose. Evaluation of the ERβ prognostic value in the comparative assessment of frequency and level of the marker expression in groups with good and poor prognosis by Ki-67 proliferative index score in breast cancer. Methods. ERβ expression level (% of cells expressing the marker) in 68 breast tissue samples was quantified by immunofluorescence and flow cytometry. Primary antibodies to ERβ (clone 14C8, ab288) and secondary antibodies conjugated with DyLight650 (ab98729) were used. In the same samples, the Ki-67 expression level was assessed by the immunohistochemical method. Results. The ERβ and Ki-67 were detected in 100% breast tissue samples with high heterogeneity of the markers’ expression in different patients. Statistical analysis of good and poor prognosis in accordance with the Ki-67 proliferative index score (Ki-67≤20% and Ki-67&gt;20%) showed the prognostic value of the ERβ expression level of 50%. There was no association between the Ki-67 and ERβ expression levels in the same tumor sample (Spearman's rank correlation coefficient R=–0,16; P&gt;0,05). At the same time, high expression of ERβ≥50% was 2,3 times more frequently detected in the good vs poor prognostic group by Ki-67 — 41% vs 18%, P=0,02. Conclusion. The ERβ expression level ≥50% in the tumor can be considered as a factor of good prognosis of breast cancer.

WTAP-mediated m6A modification on circCMTM3 inhibits hepatocellular carcinoma ferroptosis by recruiting IGF2BP1 to increase PARK7 stability.

Hepatocellular carcinoma (HCC) has poor prognosis and high mortality. CircCMTM3 was significantly up-regulated in HCC. However, the mechanism of circCMTM3 in HCC is not full elucidated. The expression level of circCMTM3, PARK7, GPX4, and Ki67 in HCC cells and tissues were quantified by qRT-PCR, IHC, and Western blotting. The level of GSH, total iron, Fe2+, and MDA were detected by their kits. CCK-8 and flow cytometry analysis were used to evaluated cell proliferation and lipid ROS level, respectively. m6A level of circCMTM3 was assessed by MeRIP-PCR. RNA pulldown, RIP, and FISH detected the interaction between circCMTM3, WTAP, and PARK7. Tumor xenograft model was constructed to validate the function of cicrCMTM3 and WTAP. CircCMTM3 and WTAP were enhanced in HCC tissues and cells. Knockdown of WTAP inhibited m6A modification of circCMTM3, which promoted HCC ferroptosis. circCMTM3 silencing suppressed the expression and stability of PARK7 through binding with IGF2BP1 in HCC cells, which finally induced ferroptosis. In vivo studies demonstrated that silencing WTAP and circCMTM3 suppressed tumor growth and promoted HCC ferroptosis in nude mice by regulating PARK7 signaling. CircCMTM3 promoted the carcinogenesis through inhibiting ferroptosis by recruiting IGF2BP1 to increase PARK7 stability in HCC, suggesting that cicrCMTM3 may be an important marker for HCC treatment.

Chloroform Fraction of Drymaria cordata Linn (CFDC) Suppresses Estradiol Benzoate- Induced Endometrial Hyperplasia

The diagnosis of uterine dysfunction (endometrial hyperplasia) is on the rise. The available treatment is quite expensive and associated with some side effects. The therapeutic potential of natural products is now being explored, as they are easily available with little or no side effects. Drymaraia cordata is folklorically utilized in the treatment of diverse ailments including uterine fibroids. This study aims to investigate the potential therapeutic effect of chloroform fraction of methanol extract of Drymaria cordata (CFDC) in estradiol benzoate (EB)-induced endometrial hyperplasia. Thirty-six rats were randomly divided equally into six groups. These included control group, CFDC: (100 mg/kg), CFDC: (200 mg/kg), EB: (2 mg/kg), EB + CFDC (100 mg/kg), and EB + CFDC (200 mg/kg). Endometrial hyperplasia (EH) was induced by intraperitoneal injection of EB. The levels of estrogen (E2), progesterone (PG), Follicle stimulating hormone (FSH), Luteinizing hormone (LH), Malondialdehyde (MDA), Superoxide dismutase (SOD), and Glutathione peroxidase (GSH-Px) activities were determined using ELISA technique. The uterine histological assessment and immunohistochemical expression levels of estrogen receptor, Ki-67, cytochrome c, and caspase 3 were carried out. EH was severely expressed in the uterine section of EB-treated rats. However, CFDC administration improved the pathological features of the animal model. The sex hormones levels were increased in the EB-treated group, which were significantly reduced by CFDC. The antioxidant indices were also restored by CFDC. Immunoexpression levels of ERα and Ki-67 were downregulated while cytochrome c and caspase 3 were upregulated by CFDC. This study suggests that CFDC contains phytochemicals that can protect against EB-induced EH via modulation of hormonal signaling, apoptotic machinery, and oxidative indices.