In vivo and in vitro studies on the role of regulating Smac expression in the occurrence and development of colon cancer.

We aimed to explore the role of regulating Smac expression levels in the occurrence and development of colon cancer through in vitro and in vivo experiments. Colon cancer cells HT-29 were cultured and transfected into different groups. qRT-PCR was used to detect the expression level of Smac in cells; Flow cytometry was used to detect the apoptotic ability of each group of cells; Western blot was used to detect the protein expression of Smac and apoptosis-related factors Survivin and Caspase-3; The nude mouse tumorigenesis experiment was conducted to detect the regulatory effect of regulating Smac expression levels on the growth of colon cancer transplanted tumors in vivo. In comparison to the FHC group, the HT-29 group exhibited a decrease in Smac expression. The si-Smac group, when compared with the si-NC group, showed significant reductions in Smac mRNA and protein levels, weaker cell apoptosis, increased Survivin, and decreased Caspase-3 expression. Contrarily, the oe-Smac group, against the oe-NC group, displayed increased Smac mRNA and protein levels, enhanced apoptosis, reduced Survivin, and elevated Caspase-3 expression. In nude mice tumor transplantation experiments, the LV-sh-Smac group, as opposed to the LV-sh-NC group, had tumors with greater volume and weight, reduced Smac and Caspase-3, and increased Survivin expression. In contrast, the LV-oe-Smac group, compared with the LV-oe-NC group, showed tumors with decreased volume and mass, increased expressions of Smac and Caspase-3, and decreased Survivin. Smac is lowly expressed in colon cancer. Upregulation of Smac expression can inhibit the occurrence and development of colon cancer, possibly by inhibiting Survivin expression and promoting Caspase-3 expression, thereby enhancing the pro-apoptotic function.

Increasing the radiation-induced cytotoxicity by silver nanoparticles and docetaxel in prostate cancer cells.

BACKGROUND: Radiation therapy is utilized for treatment of localized prostate cancer. Nevertheless, cancerous cells frequently develop radiation resistance. While higher radiation doses have not always been effective, radiosensitizers have been extensively studied for their ability to enhance the cytotoxic effects of radiation. So, this study aims to evaluate the possible radiosensitization effects of docetaxel (DTX) and silver nanoparticles (SNP) in LNCaP cells. METHODS: The cytotoxic effects of DTX, SNP and 2 Gy of X-Ray radiation treatments were assessed in human LNCaP cell line using the MTT test after 24 h. Moreover, the effects of DTX, SNP and radiation on Epidermal growth factor (EGF), Caspase 3, inducible nitric oxide synthase and E-cadherin gene expression were analyzed using the Real-time PCR method. The level of Hydrogen peroxide (H2O2), an oxidative stress marker, was also detected 24 h after various single and combined treatments. RESULTS: The combinations of SNP (in low toxic concentration) and/or DTX (0.25× IC50 and 0.5 × IC50 concentrations for triple and double combinations respectively) with radiation induced significant cytotoxicity in LNCaP cells in comparison to monotherapies. These cytotoxic effects were associated with the downregulation of EGF mRNA. Additionally, H2O2 levels increased after Radiation + SNP + DTX triple combination and double combinations including Radiation + SNP and Radiation + DTX versus single treatments. The triple combination treatment also increased Caspase 3 and and E-cadherin mRNA levels in compared to single treatments in LNCaP cells. CONCLUSION: Our results indicate that the combination of SNP and DTX with radiation induces significant anti-cancer effects. Upregulation of Caspase 3 and E-cadherin gene expression, and decreased mRNA expression level of EGF may be exerted specifically by use of this combination versus single treatments.

Heat Shock Protein HSP27 and the Status of the Glutathione System in Dexamethasone-Induced Apoptosis of Jurkat Tumor Cells.

We studied the effect of the HSP27 inhibitor, 5-(5-ethyl-2-hydroxy-4-methoxyphenyl)-4-(4-methoxyphenyl)-isoxazole, at a final concentration of 0.1 µM and/or the apoptosis inducer dexamethasone at a final concentration of 10 µM on the content of hydroxyl radical, reduced and oxidized glutathione, HSP27, activity of glutathione reductase, glutathione peroxidase, caspase-3, and the number of Annexin+ Jurkat tumor cells. The involvement of HSP27 in apoptosis of Jurkat tumor cells was demonstrated. Simultaneous exposure to the HSP27 inhibitor and dexamethasone resulted in an increase in the level of HSP27 against the background of developing oxidative stress (increase in the concentration of hydroxyl radicals and changes in the state of the glutathione system).

Effect of Pyridoxine Derivative B6NO on Transcription Factor Nrf2 Activity and Cytotoxic Properties of Doxorubicin In Vitro.

The effect of a new pyridoxine derivative B6NO on doxorubicin cytotoxicity and Nrf2-dependent cellular processes in vitro was studied. Antioxidant B6NO enhances the cytotoxic effect of doxorubicin on tumor cells, which is associated with G2/M cell division arrest and an increase in activity of proapoptotic enzyme caspase-3. The antioxidant promotes intracellular accumulation and nuclear translocation of Nrf2 transcription factor in non-tumor and tumor cells. In non-tumor cells, B6NO increases the expression of antioxidant system proteins and reduces ROS generation in the presence of doxorubicin. In tumor cells, no activation of Nrf2-dependent processes occurs under the action of the antioxidant. Our findings demonstrate the prospect of further studies of pyridoxine derivatives as antioxidants to reduce adverse reactions during chemotherapy.

Prognostic and chemotherapeutic implications of a novel four-gene pyroptosis model in head and neck squamous cell carcinoma.

Background: Head and neck squamous cell carcinoma (HNSCC) is one of the most common cancers. Chemotherapy remains one dominant therapeutic strategy, while a substantial proportion of patients may develop chemotherapeutic resistance; therefore, it is particularly significant to identify the patients who could achieve maximum benefits from chemotherapy. Presently, four pyroptosis genes are reported to correlate with the chemotherapeutic response or prognosis of HNSCC, while no study has assessed the combinatorial predicting efficacy of these four genes. Hence, this study aims to evaluate the predictive value of a multi-gene pyroptosis model regarding the prognosis and chemotherapeutic responsiveness in HNSCC. Methods: By utilizing RNA-sequencing data from The Cancer Genome Atlas database and the Gene Expression Omnibus database, the pyroptosis-related gene score (PRGscore) was computed for each HNSCC sample by performing a Gene Set Variation Analysis (GSVA) based on four genes (Caspase-1, Caspase-3, Gasdermin D, Gasdermin E). The prognostic significance of the PRGscore was assessed through Cox regression and Kaplan-Meier survival analyses. Additionally, chemotherapy sensitivity stratified by high and low PRGscore was examined to determine the potential association between pyroptosis activity and chemosensitivity. Furthermore, chemotherapy sensitivity assays were conducted in HNSCC cell lines in vitro. Results: As a result, our study successfully formulated a PRGscore reflective of pyroptotic activity in HNSCC. Higher PRGscore correlates with worse prognosis. However, patients with higher PRGscore were remarkably more responsive to chemotherapy. In agreement, chemotherapy sensitivity tests on HNSCC cell lines indicated a positive association between overall pyroptosis levels and chemosensitivity to cisplatin and 5-fluorouracil; in addition, patients with higher PRGscore may benefit from the immunotherapy. Overall, our study suggests that HNSCC patients with higher PRGscore, though may have a less favorable prognosis, chemotherapy and immunotherapy may exhibit better benefits in this population.

Overexpression of CISD2 alleviates septic acute kidney injury via activating Sonic Hedgehog signaling pathway.

Patients with sepsis are often complicated by acute kidney injury (AKI), which greatly increases mortality. In this study, our purpose was to explore the expression and function of CDGSH iron sulfur domain 2 (CISD2) in septic AKI, and the underlying molecular mechanism. Western blot and quantitative real-time polymerase chain reaction (RT-PCR) were employed to detect protein and mRNA levels in cells. The inflammation level of cells was evaluated by detecting the content of inflammatory factors (TNF-α, IL-1ß, IL-6). Apoptosis of cells was evaluated by Caspase-3 activity assay, flow cytometry and terminal deoxynucleotidyl transferase-mediated dUTP Nick-End Labeling (TUNEL) staining. CISD2 was down-regulated in HK-2 cells treated with lipopolysaccharide (LPS). LPS treatment increased the level of inflammatory factors, the activity of Caspase-3, and the rate of apoptosis in HK-2 cells. However, overexpression of CISD2 significantly suppressed these effects. Moreover, overexpression of CISD2 activated the Sonic Hedgehog (SHH) signaling pathway. The use of cyclopamine (Cyc), a SHH signaling pathway inhibitor, eliminated the effect of overexpressing CISD2, that is, inhibiting LPS-induced inflammation and apoptosis of HK-2 cells. LPS treatment down-regulated CISD2 in HK-2 cells, and overexpression of CISD2 could inhibit LPS-induced inflammation and apoptosis of HK-2 cells by activating the SHH signaling pathway.

Erythropoietin alleviates lung ischemia-reperfusion injury by activating the FGF23/FGFR4/ERK signaling pathway.

Background: The purpose of the present study was to investigate the effect of erythropoietin (EPO) on lung ischemia-reperfusion injury (LIRI). Methods: Sprague Dawley rats and BEAS-2B cells were employed to construct an ischemia-reperfusion (I/R)-induced model in vivo and in vitro, respectively. Afterward, I/R rats and tert-butyl hydroperoxide (TBHP)-induced cells were treated with different concentrations of EPO. Furthermore, 40 patients with LIRI and healthy controls were enrolled in the study. Results: It was observed that lung tissue damage, cell apoptosis and the expression of BAX and caspase-3 were higher in the LIRI model in vivo and in vitro than in the control group, nevertheless, the Bcl-2, FGF23 and FGFR4 expression level was lower than in the control group. EPO administration significantly reduced lung tissue damage and cell apoptosis while also up-regulating the expression of FGF23 and FGFR4. Rescue experiments indicated that EPO exerted a protective role associated with the FGF23/FGFR4/p-ERK1/2 signal pathway. Notably, the expression of serum EPO, FGF23, FGFR4 and Bcl-2 was decreased in patients with LIRI, while the expression of caspase-3 and BAX was higher. Conclusion: EPO could effectively improve LIRI, which might be related to the activation of the FGF23/FGFR4/p-ERK1/2 signaling pathway.

Determination of anti-cancer effects of Nigella sativa seed oil on MCF7 breast and AGS gastric cancer cells.

BACKGROUND: This study aimed to investigate the cytotoxic, apoptotic, invasion, metastasis, and heat shock proteins (HSPs) effects of N. sativa oil on breast and gastric cancer cells. METHODS: We assessed the cytotoxic and apoptotic effects of various concentrations of N. sativa oil (10-50-100-200 µg/mL) on MCF7 breast cancer and AGS, an adenocarcinoma of the gastric cell line, at 24, 48 and 72 h using the MTT test. Additionally, the expression of the Caspase-3, BCL2/Bax, MMP2-9 and HSP60-70 gene was examined using RT-PCR in cell lines treating with N. sativa. RESULTS: The MTT experiments demonstrate that N. sativa has a time and dose-dependent inhibitory effect on the proliferation of MCF7 and AGS cancer cells. The vitality rates of MCF7 and AGS cells treated with N. sativa were 77.04-67.50% at 24 h, 65.28-39.14% at 48 h, and 48.95-32.31% at 72 h. The doses of 100 and 200 µg/mL were shown to be the most effective on both cancer cells. RT-PCR analysis revealed that N. sativa oil extract increased caspase-3 levels in both cell lines at higher concentrations and suppressed BCL2/Bax levels. Exposure of MCF7 and AGS cell lines to N. sativa caused a significant decrease in the expression of MMP2-9 and HSP60-70 genes over time, particularly at a dosage of 200 µg/mL compared to the control group (p < 0.05). CONCLUSIONS: Our findings indicate that N. sativa oil has a dose-dependent effect on cytotoxicity and the expression of apoptotic, heat shock proteins, and matrix metalloproteinases genes in breast and gastric cancer.

Methylmercury Induces Apoptosis in Mouse C17.2 Neural Stem Cells through the Induction of OSGIN1 Expression by NRF2.

Methylmercury is a known environmental pollutant that exhibits severe neurotoxic effects. However, the mechanism by which methylmercury causes neurotoxicity remains unclear. To date, we have found that oxidative stress-induced growth inhibitor 1 (OSGIN1), which is induced by oxidative stress and DNA damage, is also induced by methylmercury. Therefore, in this study, we investigated the relationship between methylmercury toxicity and the induction of OSGIN1 expression using C17.2 cells, which are mouse brain neural stem cells. Methylmercury increased both OSGIN1 mRNA and protein levels in a time- and concentration-dependent manner. Moreover, these increases were almost entirely canceled out by pretreatment with actinomycin D, a transcription inhibitor. Furthermore, similar results were obtained from cells in which expression of the transcription factor nuclear factor erythroid 2-related factor 2 (NRF2) was suppressed, indicating that methylmercury induces OSGIN1 expression via NRF2. Methylmercury causes neuronal cell death by inducing apoptosis. Therefore, we next investigated the role of OSGIN1 in methylmercury-induced neuronal cell death using the activation of caspase-3, which is involved in apoptosis induction, as an indicator. As a result, the increase in cleaved caspase-3 (activated form) induced by methylmercury exposure was decreased by suppressing OSGIN1, and the overexpression of OSGIN1 further promoted the increase in cleaved caspase-3 caused by methylmercury. These results suggest, for the first time, that OSGIN1 is a novel factor involved in methylmercury toxicity, and methylmercury induces apoptosis in C17.2 cells through the induction of OSGIN1 expression by NRF2.

Okadaic acid enhances NfKB, TLR-4, caspase 3, ERK ½, c-FOS, and 8-OHdG signaling pathways activation in brain tissues of zebrafish larvae.

This study was designed to investigate the potential neuronal damage mechanism of the okadaic acid (OA) in the brain tissues of zebrafish embryos by evaluating in terms of immunofluorescence of Nf KB, TLR-4, caspase 3, ERK ½, c-FOS and 8-OHdG signaling pathways. We also evaluated body malformations. For this purpose, zebrafish embryos were exposed to 0.5 µg/ml, 1 µg/ml and 2.5 µg/ml of OA for 5 days. After application, FITC/GFP labeled protein-specific antibodies were used in immunofluorescence assay for NfKB, TLR-4, caspase 3, ERK ½, c-FOS and 8-OHdG respectively. The results indicated that OA caused immunofluorescence positivity of NfKB, TLR-4, caspase 3, ERK ½, c-FOS and 8-OHdG in a dose-dependent manner in the brain tissues of zebrafish embryos. Pericardial edema (PE), nutrient sac edema (YSE) and body malformations, tail malformation, short tail and head malformation (BM) were detected in zebrafish embryos. These results suggest that OA induces neuronal damage by affecting the modulation of DNA damage, apoptotic, and inflammatory activities in the brain tissues of zebrafish embryos. The increase in signaling pathways shows that OA can cause damage in the structure and function of brain nerve cells. Our results provide a new basis for the comprehensive assessment of the neural damage of OA and will offer enable us to better understand molecular the mechanisms underlying the pathophysiology of OA toxicity.

Ssc-miR-7139-3p suppresses foot-and-mouth disease virus replication by promoting degradation of 3Cpro through targeting apoptosis-negative regulatory gene Bcl-2.

Foot-and-mouth disease is a highly contagious and infectious disease affecting cloven-hoofed animals. Disease control is complicated by its highly contagious nature and antigenic diversity. Host microRNAs (miRNAs) are post-transcriptional regulators that either promote or repress viral replications in virus infection. In the present study, we found that ssc-miR-7139-3p (Sus scrofa miR-7139-3p) was significantly up-regulated in host cells during foot-and-mouth disease virus (FMDV) infection. Overexpression of miR-7139-3p attenuated FMDV replication, whereas inhibition promoted FMDV replication. In addition, the survival rate of FMDV infected suckling mice was increased through injection of miR-7139-3p agomiR. Further studies revealed that miR-7139-3p targets Bcl-2 to initiate the apoptotic pathway and caspase-3 cleaved 3Cpro behind the 174th aspartic acid (D174), which eventually promotes the degradation of 3Cpro. Overall, our findings demonstrate that miR-7139-3p suppresses FMDV replication by promoting degradation of 3Cpro through targeting the apoptosis-negative regulatory gene Bcl-2.

Biochemical and molecular-level effects of co-exposure to chlorpyrifos and lambda-cyhalothrin on the earthworm (Eisenia fetida).

Farmland soil organisms frequently encounter pesticide mixtures presented in their living environment. However, the underlying toxic mechanisms employed by soil animals to cope with such combined pollution have yet to be explored. This investigation aimed to reveal the changes in cellular and mRNA levels under chlorpyrifos (CPF) and lambda-cyhalothrin (LCT) co-exposures in earthworms (Eisenia fetida). Results exhibited that the combination of CPF and LCT triggered an acute synergistic influence on the animals. Most exposures resulted in significant alterations in the activities of total superoxide dismutase (T-SOD), copper/zinc superoxide dismutase (Cu/Zn-SOD), caspase 3, and carboxylesterase (CarE) compared to the basal level. Moreover, when exposed to chemical mixtures, the transcription levels of four genes [heat shock protein 70 (hsp70), gst, sod, and calreticulin (crt)] also displayed more pronounced changes compared with their individual exposures. These changes in determined parameters indicated the occurrence of oxidative stress, cell death, detoxification dysfunction, and endoplasmic reticulum damage after co-exposure to CPF and LCT in E. fetida. The comprehensive examination of mixture toxicities of CPF and LCT at different endpoints would help to understand the overall toxicity they cause to soil invertebrates. The augmented deleterious effect of these pesticides in a mixture suggested that mixture toxicity assessment was necessary for the safety evaluation and application of pesticide mixtures.

MiR-98-5p plays suppressive effects on IL-1ß-induced chondrocyte injury associated with osteoarthritis by targeting CASP3.

BACKGROUND: This study aims to explore how miR-98-5p affects osteoarthritis, focusing on its role in chondrocyte inflammation, apoptosis, and extracellular matrix (ECM) degradation. METHODS: Quantitative real-time PCR was used to measure miR-98-5p and CASP3 mRNA levels in OA cartilage tissues and IL-1ß-treated CHON-001 cells. We predicted miR-98-5p and CASP3 binding sites using TargetScan and confirmed them via luciferase reporter assays. Chondrocyte viability was analyzed using CCK-8 assays, while pro-inflammatory cytokines (IL-1ß, IL-6, TNF-α) were quantified via ELISA. Caspase-3 activity was examined to assess apoptosis, and Western blotting was conducted for protein marker quantification. RESULTS: Our results showed lower miR-98-5p levels in both OA cartilage and IL-1ß-stimulated cells. Increasing miR-98-5p resulted in reduced pro-inflammatory cytokines, decreased caspase-3 activity, and improved cell viability. Furthermore, miR-98-5p overexpression hindered IL-1ß-induced ECM degradation, evident from the decline in MMP-13 and ß-catenin levels, and an increase in COL2A1 expression. MiR-98-5p's impact on CASP3 mRNA directly influenced its expression. Mimicking miR-98-5p's effects, CASP3 knockdown also inhibited IL-1ß-induced inflammation, apoptosis, and ECM degradation. In contrast, CASP3 overexpression negated the suppressive effects of miR-98-5p. CONCLUSIONS: In conclusion, our data collectively suggest that miR-98-5p plays a protective role against IL-1ß-induced damage in chondrocytes by targeting CASP3, highlighting its potential as a therapeutic target for OA.

[miR-515-5p targeting Toll-like receptor 4 regulates myeloid differentiation primary response gene 88/nuclear factor-kappa B pathway to inhibit apoptosis and inflammatory response of osteoarthritis chondrocytes].

Objective: To explore the molecular mechanism of miR-515-5p in inhibiting chondrocyte apoptosis and alleviating inflammatory response in osteoarthritis (OA). Methods: Human cartilage cell line C28/I2 was cultured in vitro and treated with 10 ng/mL interleukin 1ß (IL-1ß) for 24 hours to construct an in vitro OA model. C28/I2 cells were transfected with miR mimics, mimics negative control (NC), over expression (oe)-NC, and oe-Toll-like receptor 4 (TLR4), respectively, and then treated with 10 ng/mL IL-1ß for 24 hours to establish OA model. Cell proliferation capacity was detected by cell counting kit 8 and 5-Ethynyl-2'-deoxyuridine, cell apoptosis and cell cycle were detected by flow cytometry, and B-cell lymphoma 2 protion (Bcl-2), Bcl-2-associated X protein (Bax), cleaved-Caspase-3, TLR4, myeloid differentiation primary response gene 88 (MyD88), p65 and phosphorylated p65 (p-p65) protein expression levels were detected by Western blot. Real-time fluorescence quantitative PCR was used to detect mRNA expression levels of miR-515-5p and TLR4, and ELISA was used to detect pro-inflammatory factor prostaglandin E2 (PGE2), tumor necrosis factor α (TNF -α), and IL-6 levels in cell supernatant. The potential binding sites between miR-515-5p and TLR4 were predicted by BiBiServ2 database, and the targeting relationship between miR-515-5p and TLR4 was verified by dual luciferase reporting assay. Results: After the treatment of C28/I2 cells with IL-1ß, the expressions of miR-515-5p and Bcl-2 protein and the proliferation ability of C28/I2 cells significantly reduced. The expression levels of Bax and cleaved-Caspase-3 protein, the levels of pro-inflammatory factors (PGE2, TNF-α, IL-6) in the supernatant of C28/I2 cells, and the apoptosis of C28/I2 cells significantly increased. In addition, the proportion of the cells at S phase and G 2 phase decreased significantly, and the proportion of cells at G 1 phase increased significantly, suggesting that the cell cycle was blocked after IL-1ß treatment. After transfection with miR mimics, the expression level of miR-515-5p in the cells significantly up-regulated, partially reversing the apoptosis of OA chondrocytes induced by IL-1ß, and alleviating the cycle arrest and inflammatory response of OA chondrocytes. After treating C28/I2 cells with IL-1ß, the mRNA and protein levels of TLR4 significantly increased. Overexpression of miR-515-5p targeted inhibition of TLR4 expression and blocked activation of MyD88/nuclear factor κB (NF-κB) pathway. Overexpression of TLR4 could partially reverse the effect of miR mimics on IL-1ß-induced apoptosis and inflammation of OA chondrocytes. Conclusion: miR-515-5p negatively regulates the expression of TLR4, inhibits the activation of MyD88/NF-κB pathway and apoptosis of OA chondrocytes, and effectively alleviates the inflammatory response of the cells.

Thymoquinone potentiates anti-cancer effects of cisplatin in oral squamous cell carcinoma via targeting oxidative stress.

Recent evidence has proved that thymoquinone as a natural polyphenol has great anticancer and anti-proliferative effects in cancer cells. In this study, we aimed to examine the effects of thymoquinone on increasing cisplatin-induced apoptosis human oral squamous cell carcinoma cells and its underlying molecular mechanisms. SCC-25 cancer cells treated by thymoquinone and cisplatin with different concentrations. Cell viability will determine by using MTT assay. The concentrations of reactive oxygen species (ROS) and antioxidant activities were determined using specific related kits. DNA damage, lipid, and protein oxidation were assessed. Real-time PCR and Western blot analysis will be used to determine the expression of apoptosis-related proteins including Bax, Bcl-2, and caspase-3. Combination of thymoquinone and cisplatin suppressed synergistically SCC-25 cancer cell viability and induced apoptosis in dose-depended manner. Cell treatment with combination of thymoquinone and cisplatin led to accumulation of ROS within cells and increase in the intracellular levels of DNA damage, protein and lipid peroxidation. In addition, the combination of thymoquinone and cisplatin modulated the mRNA and protein expression levels of apoptosis-related proteins including Bax, Bcl-2, and caspase-3. Thymoquinone potentiated cisplatin anti-cancer effect on OSCC by inducing oxidative stress in cells.

Cytoprotective role of human dental pulp stem cell-conditioned medium in chemotherapy-induced alopecia.

BACKGROUND: Chemotherapy-induced alopecia (CIA) is a distressing adverse effect of chemotherapy, with an estimated incidence of 65% and limited treatment options. Cyclophosphamide (CYP) is a common alopecia-inducing chemotherapy agent. Human dental pulp stem cells (DPSCs) secrete several paracrine factors that up-regulate hair growth. Conditioned medium (CM) collected from DPSCs (DPSC-CM) promotes hair growth; culturing mesenchymal stem cells under hypoxic conditions can enhance this effect. METHODS: The effect of DPSC-CM cultured under normoxic (N-) and hypoxic (H-) conditions against CYP-mediated cytotoxicity in keratinocytes was examined using cell viability assay, lactate dehydrogenase (LDH) cytotoxicity assay, and apoptosis detection. The damage-response pathway was determined in a well-established CIA mouse model by analyzing macroscopic effects, histology, and apoptosis. Reverse transcription-quantitative PCR and Caspase-3/7 activity assay were used to investigate the impact of DPSC-CM on the molecular damage-response pathways in CYP-treated mice. The effect of post-CIA DPSC-CM application on post-CIA hair regrowth was analyzed by macroscopic effects and microstructure observation of the hair surface. Furthermore, to investigate the safety of DPSC-CM as a viable treatment option, the effect of DPSC-CM on carcinoma cell lines was examined by cell viability assay and a subcutaneous tumor model. RESULTS: In the cell viability assay, DPSC-CM was observed to increase the number of keratinocytes over varying CYP concentrations. Furthermore, it reduced the LDH activity level and suppressed apoptosis in CYP-treated keratinocytes. DPSC-CM exhibited the cytoprotective role in vivo via the dystrophic anagen damage-response pathway. While both N-CM and H-CM downregulated the Caspase-3/7 activity level, H-CM downregulated Caspase-3 mRNA expression. The proportion of post-CIA H-CM-treated mice with > 90% normal hair was nearly twice that of vehicle- or N-CM-treated mice between days 50 and 59 post-depilation, suggesting that post-CIA H-CM application may accelerate hair regrowth and improve hair quality. Furthermore, DPSC-CM suppressed proliferation in vitro in certain carcinoma cell lines and did not promote the squamous cell carcinoma (SCC-VII) tumor growth rate in mice. CONCLUSIONS: The potentiality of DPSC-CM and H-CM as a promising cytoprotective agent and hair regrowth stimulant, respectively, for CIA needs in-depth exploration.

Borax induces ferroptosis of glioblastoma by targeting HSPA5/NRF2/GPx4/GSH pathways.

Glioblastoma multiforme (GBM) is a highly aggressive and lethal form of primary brain tumour. Borax has been demonstrated to exhibit anti-cancer activity through cell death pathways. However, the specific impact of borax on ferroptosis in GBM is not well-established, and the underlying regulatory mechanisms remain unclear. Initially, the effective concentration of borax on cell viability and proliferation in U251 and A172 cells was determined. Subsequently, the effects of borax on the wound healing were analysed. Nuclear factor erythroid 2-related factor 2 (NRF2), glutathione peroxidase 4 (GPx4), glutathione (GSH), HSP70 protein 5 (HSPA5), malondialdehyde (MDA) levels and caspase-3/7 activity were determined in borax-treated and untreated cells. Finally, the protein expression levels of HSPA5, NRF2 and GPx4 were analysed. Borax suppressed cell viability and proliferation in U251 and A172 cells in a concentration- and time-dependent manner. In addition, borax treatment decreased GPx4, GSH, HSPA5 and NRF2 levels in U251 and A172 cells while increasing MDA levels and caspase-3/7 activity. Moreover, borax reduced mRNA and protein levels of HSPA5, NRF2 and GPx4 in U251 and A172 cells. Consequently, borax may induce ferroptosis in GBM cells and regulate the associated regulatory mechanisms targeting NRF2 and HSPA5 pathways. This knowledge may contribute to the development of novel therapeutic approaches targeting ferroptosis in GBM and potentially improve patient outcomes.

Derivation of a novel antimicrobial peptide from the Red Sea Brine Pools modified to enhance its anticancer activity against U2OS cells.

Cancer associated drug resistance is a major cause for cancer aggravation, particularly as conventional therapies have presented limited efficiency, low specificity, resulting in long term deleterious side effects. Peptide based drugs have emerged as potential alternative cancer treatment tools due to their selectivity, ease of design and synthesis, safety profile, and low cost of manufacturing. In this study, we utilized the Red Sea metagenomics database, generated during AUC/KAUST Red Sea microbiome project, to derive a viable anticancer peptide (ACP). We generated a set of peptide hits from our library that shared similar composition to ACPs. A peptide with a homeodomain was selected, modified to improve its anticancer properties, verified to maintain high anticancer properties, and processed for further in-silico prediction of structure and function. The peptide's anticancer properties were then assessed in vitro on osteosarcoma U2OS cells, through cytotoxicity assay (MTT assay), scratch-wound healing assay, apoptosis/necrosis detection assay (Annexin/PI assay), RNA expression analysis of Caspase 3, KI67 and Survivin, and protein expression of PARP1. L929 mouse fibroblasts were also assessed for cytotoxicity treatment. In addition, the antimicrobial activity of the peptide was also examined on E coli and S. aureus, as sample representative species of the human bacterial microbiome, by examining viability, disk diffusion, morphological assessment, and hemolytic analysis. We observed a dose dependent cytotoxic response from peptide treatment of U2OS, with a higher tolerance in L929s. Wound closure was debilitated in cells exposed to the peptide, while annexin fluorescent imaging suggested peptide treatment caused apoptosis as a major mode of cell death. Caspase 3 gene expression was not altered, while KI67 and Survivin were both downregulated in peptide treated cells. Additionally, PARP-1 protein analysis showed a decrease in expression with peptide exposure. The peptide exhibited minimal antimicrobial activity on critical human microbiome species E. coli and S. aureus, with a low inhibition rate, maintenance of structural morphology and minimal hemolytic impact. These findings suggest our novel peptide displayed preliminary ACP properties against U2OS cells, through limited specificity, while triggering apoptosis as a primary mode of cell death and while having minimal impact on the microbiological species E. coli and S. aureus.

CRISPR du-HITI an attractive approach to targeting Long Noncoding RNA HCP5 as inhibitory factor for proliferation of ovarian cancer cell.

This research provides a glimmer of hope that the knockout of HCP5 leads to a therapy response to considerably prolong the life of patients with OC. RT-PCR evaluated the expression of lncRNA HCP5 in the ovarian cancer OVCAR-3 cell line. CRISPR knockout cell lines validated by western blot. Small genomic deletions at the targeted locus were induced. CCK-8 colony formation assays were used to analyze the effect of HCP5 knockout on the proliferation capacity of OVCAR-3 cells. Transwell migration and invasion assayed. Furthermore, the Sphere-formation assay isolated the most aggressive population of cancer stem cells. Bioinformatic analysis showed a significant correlation between lncRNA HCP5 up-regulation and OVCAR-3 cell proliferation. The ChIP technique assesses specific sites of interaction between transcription factors and DNA. Real-time PCR assays explored the relationship between HCP5, Hsa-miR-9-5p, CXCR4, CDH1, caspase-3, p53, bcl2 and survivin. PCR carried out amplification of the 448-bp band for sgRNA1 and sgRNA2 after the use of particular primers for HCP5. the number of breast cancer cells that moved to the bottom chamber reduced considerably after transfection with PX461-sgRNA1/2 vectors compared to the Blank control groups (P < 0.05). MTT assay designated growth curves that showed the rate of OVCAR-3 growth was significantly repressed (***P < 0.001) when compared with control OVCAR-3 cells after HCP5 knockdown. Also, the survival results of W.T cells in 24, 48 and 72 h showed 92%, 87% and 85%, respectively. This is while the cells of the CRISPR/Cas9 group in which LncRNA HCP5 was knocked out had 42% (*P < 0.05), 23%(**P < 0.01) and 14% (**P < 0.01) survival, respectively. The expression levels of caspase-3, Hsa-miR-9-5p, P53 genes in the HCP5 deletion of CRISPR/Cas9 group significantly increased than the W.T. control group; the deletion group showed a considerable reduction in HCP5 expression compared to the blank control group (3.6-fold, p < 0.01). Whereas BCL2, SURVIVIN, CXCR4, CDH1 genes expression markedly increased than in HCP5 knockout cells (5.8-fold, p < 0.05). These results indicate that CRISPR/Cas9-mediated HCP5 disruption on OVCAR-3 cell lines promotes anti-tumor biomarkers, suppressing ovarian cancer progression. Consistent with these results, HCP5 is one of the most critical lnc for the efficient proliferation and migration of OVCAR-3 cell lines.

PEI, a new transfection method, augments the inhibitory effect of RBM5 on prostate cancer.

PEI is a cationic polymer, serving as a non-viral transfection carrier grounded in nanotechnology that enhances transfection efficiency via the proton sponge effect. RBM5 is an RNA-binding protein that can inhibit tumor development. This study involved the transfection of RBM5 in prostate cancer cells with PEI, Lipo2000, and their combination. Transwell and wound healing assays were used to observe invasion and migration of prostate cancer cells and flow cytometry was used to observe the apoptosis. Detect the expression of invasion and migration-related protein MMP9 through western blotting experiment. An activity detection kit was used to detect the activity of apoptotic protein caspase-3. We found that there was no significant difference in transfection efficiency when PEI and Lipo2000 were used alone but it significantly improved when they are combined. RBM5 reduced invasion, migration, and proliferation of prostate cancer and enhanced apoptosis. MMP9 expression was reduced, and the activity of caspase-3 was increased. PEI transfection could improve the inhibition of RBM5 on tumors more than Lipo2000. The inhibitory effect is more obvious when the two are used together. RBM5 transfected with PEI can amplify its inhibitory effect on prostate cancer, and this effect is more evident when combined with Lipo2000.