Search Results (657)

Recurrent pregnancy loss (RPL) is defined as the occurrence of two or more pregnancy losses before 20 weeks of gestation. RPL is a common medical condition among reproductive-age women, with approximately 23 million cases reported annually worldwide. Up to 5% of pregnant women may experience two or more consecutive pregnancy losses. Previous studies have investigated risk factors for RPL, including maternal age, uterine pathology, genetic anomalies, infectious agents, endocrine disorders, thrombophilia, and immune dysfunction. However, RPL is a disease caused by a complex interaction of genetic factors, environmental factors (e.g., diet, lifestyle, and stress), epigenetic factors, and the immune system. In addition, due to the lack of research on genetics research related to RPL, the etiology remains unclear in up to 50% of cases. Platelets play a critical role in pregnancy maintenance. This study examined the associations of platelet receptor and ligand gene variants, including integrin subunit beta 3 (ITGB3) rs2317676 A > G, rs3809865 A > T; fibrinogen gamma chain (FGG) rs1049636 T > C, rs2066865 T > C; glycoprotein 1b subunit alpha (GP1BA) rs2243093 T > C, rs6065 C > T; platelet endothelial cell adhesion molecule 1 (PECAM1) rs2812 C > T; and platelet endothelial aggregation receptor 1 (PEAR1) rs822442 C > A, rs12137505 G > A, with RPL prevalence. In total, 389 RPL patients and 375 healthy controls (all Korean women) were enrolled. Genotyping of each single nucleotide polymorphism was performed using polymerase chain reaction–restriction fragment length polymorphism and the TaqMan genotyping assay. All samples were collected with approval from the Institutional Review Board at Bundang CHA Medical Center. The ITGB3 rs3809865 A > T genotype was strongly associated with RPL prevalence (pregnancy loss [PL] ≥ 2: adjusted odds ratio [AOR] = 2.505, 95% confidence interval [CI] = 1.262–4.969, p = 0.009; PL ≥ 3: AOR = 3.255, 95% CI = 1.551–6.830, p = 0.002; PL ≥ 4: AOR = 3.613, 95% CI = 1.403–9.307, p = 0.008). The FGG rs1049636 T > C polymorphism was associated with a decreased risk in women who had three or more pregnancy losses (PL ≥ 3: AOR = 0.673, 95% CI = 0.460–0.987, p = 0.043; PL ≥ 4: AOR = 0.556, 95% CI = 0.310–0.997, p = 0.049). These findings indicate significant associations of the ITGB3 rs3809865 A > T and FGG rs1049636 T > C polymorphisms with RPL, suggesting that platelet function influences RPL in Korean women. Full article

Background/Objectives: P-glycoprotein (P-gp), an ATP-binding cassette (ABC) transporter, plays a key role in multidrug resistance by actively exporting chemotherapeutic agents and xenobiotics from cells. Overexpression of P-gp significantly reduces intracellular drug accumulation and compromises treatment efficacy. Despite extensive research, clinically approved P-gp inhibitors remain elusive due to toxicity, poor specificity, and limited efficacy. This study investigates DMH1, a selective type I BMP receptor inhibitor, as a novel P-gp inhibitor. Methods: DMH1 cytotoxicity was assessed in P-gp-overexpressing (PC3-TxR, K562/Dox) and P-gp-deficient (PC3) cell lines using MTT assays. P-gp inhibition was evaluated using calcein AM retention and daunorubicin (DNR) accumulation assays. Kinetic analysis determined DMH1’s effect on P-gp-mediated transport (Vmax and Km). ATPase activity assays were performed to assess DMH1’s impact on ATP hydrolysis. Preliminary molecular docking (CB-Dock2) was used to predict DMH1’s binding site on the human P-gp structure (PDB ID: 6QEX). Results: DMH1 showed no cytotoxicity in P-gp-overexpressing or deficient cells. It significantly enhanced intracellular accumulation of Calcein AM and DNR, indicating effective inhibition of P-gp function. Kinetic data revealed that DMH1 reduced Vmax without affecting Km, consistent with noncompetitive, allosteric inhibition. DMH1 also inhibited ATPase activity in a dose-dependent manner. Docking analysis suggested DMH1 may bind to an allosteric site in the transmembrane domain, potentially stabilizing the inward-facing conformation. Conclusions: DMH1 is a promising noncompetitive, allosteric P-gp inhibitor that enhances intracellular drug retention without cytotoxicity, supporting its potential as a lead compound to overcome multidrug resistance and improve chemotherapeutic efficacy. Full article

Obesity affects approximately 30% of pregnancies worldwide and is one of the leading metabolic disorders among pregnant women. Maternal obesity is often associated with placental dysfunction and structural alterations, which increase the risk of developing complications. Efflux transporters, including P-glycoprotein (P-gp), may impact placental function and fetal development. Consequently, our research examined the effects of obesity on P-glycoprotein expression in both a rat model and human placental tissue. P-gp expression was measured by RT-PCR and Western blot techniques in human and rat placental tissues. Moreover, we further characterized the high-fat and high-sugar diet (HFHSD)-induced gestational obesity rat model by measuring tissue weights. Significant decreases were observed in fetal, placental, and uterus weights in the obese animals near the end of pregnancy. In obese rats, mRNA and protein expression of placental P-gp showed a reduction on gestation days 15, 20, and 22. A similar P-gp reduction was observed in the term placenta in obese women in mRNA and protein levels. We hypothesize that the reduced expression of P-gp may heighten the susceptibility of both the fetus and placenta to P-gp substrates. This alteration could potentially result in an increased risk of pregnancy complications and obesity-related drug contraindications linked to P-gp transport during pregnancy. Full article

(1) Background: Overexpression of P-glycoprotein (P-gp) is one mediator of multidrug resistance in cancer. While many studies demonstrate the efficacy of modulating P-glycoprotein expression to increase drug response in cancer cells, the nature of the mathematical relationship between drug sensitivity and P-glycoprotein surface density is not yet characterized. (2) Methods: In this study, we employ siRNA to modulate P-gp expression in two model cell lines and evaluate their steady-state response to three common chemotherapeutics in vitro. Additionally, we model the kinetics of calcein-AM, a P-gp substrate, as a function of P-gp expression. (3) Results: For both cell lines, a robust linear relationship governs chemotherapeutic sensitivity as a function of P-gp expression, demonstrating that characterization of P-gp surface density is a strong indicator of drug response in drug-resistant cells. Furthermore, calcein accumulation and initial influx rate exhibit first-order kinetics with respect to P-gp density, further elucidating the nature of substrate interactions with P-gp-overexpressing cells. When transport kinetics are evaluated using a Michaelis–Menten model, V_max varies with P-gp density according to a first-order relationship. (4) Conclusions: These results establish the mathematical relationships between chemotherapeutic response and substrate influx as a function of P-gp expression and suggest that rational changes in P-gp expression could be used as a predictive measure of drug sensitivity in model cell lines. Full article

Background/Objectives: HIF-1α and ERRα are both implicated in breast cancer progression, yet their functional interplay remains poorly understood. This study investigates their molecular crosstalk in the context of hypoxia-induced drug resistance. Methods: MCF-7 (estrogen receptor, ER-positive) spheroids and CoCl₂-treated SK-BR-3 (ER-negative) cells were used to model tumor hypoxia. Protein expression, coimmunoprecipitation, chromatin immunoprecipitation (ChIP), pharmacological inhibition, and siRNA-mediated gene silencing were employed to assess physical and functional interactions. Immunohistochemistry (IHC) on a tissue microarray (TMA) of 168 invasive breast carcinomas was performed to evaluate clinical relevance. Results: ERRα levels remained unchanged under hypoxia, while its coactivator, Peroxisome Proliferator-Activated Receptor Gamma Coactivator-1 α (PGC-1α), was upregulated. ERRα physically interacted with HIF-1α and was required for HIF-1 transcriptional activity under hypoxic conditions. ChIP assays showed that ERRα-driven overexpression of Permeability glycoprotein 1 (P-gp) and Vascular Endothelial Growth Factor (VEGF) was mediated by HIF-1α binding to the MDR1 and VEGF promoters. Inhibition or silencing of ERRα reversed P-gp overexpression and restored intracellular doxorubicin. TMA analysis confirmed the clinical correlation between ERRα, HIF-1α, and P-gp expression, highlighting the role of ERRα in hypoxia-induced drug resistance. ERRα expression was independent of ER status, suggesting an estrogen-independent function. Conclusions: This study identifies a novel physical and functional interaction between ERRα and HIF-1α that promotes chemoresistance in hypoxic breast tumors. Targeting ERRα may represent a promising therapeutic strategy to overcome drug resistance in aggressive, ER-independent breast cancer subtypes. Full article

P-glycoprotein (P-gp) is a key element of cancer treatment resistance, actively extruding cytotoxic drugs from cells and diminishing their efficacy. While its role at the plasma membrane is well established, its intracellular localization, particularly on lysosomes, is increasingly recognized as a critical contributor to drug resistance. This study investigates four innovative LightSpot fluorescent compounds to detect and quantify both membrane and lysosomal P-gp in Triple-Negative Breast Cancer (TNBC) SUM1315 and DU4475 cell lines. Results highlighted lysosomal P-gp staining by the LightSpot-FL-1, LightSpot-BrX-1, and LightSpot-BdO-1 fluorescent compounds (Mander’s coefficients > 0.8 overlapping with LAMP2 immunostaining). After both cell lines were exposed to Olaparib, a significant increase in P-gp expression level and lysosomal distribution of P-gp was detected. Indeed, after 100 µM Olaparib exposure, LightSpot-FL-1 allowed us to quantify an increase in P-gp-positive lysosome number of 1293 and 334% for SUM1315 and DU4475 cells, respectively, compared to the control. Findings suggest that P-gp may relocate to lysosomes upon drug exposure, highlighting a dual resistance mechanism involving both membrane and lysosomal P-gp. This study demonstrated the potential of LightSpot fluorescent compounds to evaluate P-gp-mediated cell resistance to treatment and emphasized the need to assess global cell P-gp expression to improve cancer diagnosis. Full article

A series of novel N-Mannich bases derived from a dimethylpyridine–1,2,4-triazole hybrid was synthesized and evaluated in vitro for cytotoxic activity on several human gastrointestinal cancer cells (EPG, Caco-2, LoVo, LoVo/Dx, and HT-29). Compound 6 bearing a phenyl group at the N-4 position and a 4-methylphenyl piperazine moiety at the N-2 position of the 1,2,4-triazole-3-thione scaffold exerted good cytotoxic activities on EPG and Caco-2 cell lines, along with pronounced selectivity, showing lower cytotoxicity against normal colonic epithelial cells (CCD 841 CoTr). Further evaluation revealed the good ability of compound 6 to inhibit the efflux function of P-glycoprotein in P-gp-expressing cell lines (HT-29, LoVo, and LoVo/Dx). Moreover, compound 6 induced apoptotic cell death through a significant increase in the caspase-3 and p53 protein levels in HT-29 cells. Finally, the molecular docking method was applied to explain our experimental findings. The molecular modeling study based on Density Functional Theory (DFT) and the Quantum Theory of Atoms in Molecules (QTAIM) analysis provided insight into the geometric and electronic structure properties of the compounds. Full article

The development of resistance to standard cytostatics, such as 5-fluorouracil (5-FU), significantly limits the efficacy of colon cancer therapy, prompting the search for novel anticancer agents, particularly among natural compounds. This study evaluated the anticancer effects of isorhamnetin, a plant-derived flavonol, and its ability to modulate the expression of drug-resistance-related biomarkers in SW-480 and HT-29 colon cancer cells, with a focus on ATP-binding cassette (ABC) transporters. Isorhamnetin demonstrated strong cytotoxic and proapoptotic activity on both cell lines, while showing lower toxicity toward normal HaCaT cells. In addition to suppressing the mRNA expression of drug-metabolizing enzymes (CYP1A1 and CYP1B1), isorhamnetin significantly reduced the mRNA levels of multidrug resistance-associated proteins 1 and 5 (MRP1 and MRP5), as well as the P-glycoprotein (P-gp) level in SW-480 and HT-29 cells. Molecular docking analysis revealed a high binding affinity of isorhamnetin to CYP1A1, CYP1B1, P-gp, MRP1, MRP5, and glutathione S-transferase (GST) proteins, with stronger interactions than those observed for 5-FU, suggesting potential interference with their function. These results provide a solid basis for future investigations to confirm the therapeutic potential of isorhamnetin as a modulator of drug resistance in colon cancer cells. Full article

Background/Objectives: Baculoviruses represent promising gene delivery vectors for mammalian systems, combining high safety profiles with substantial cargo capacity. While pseudotyping with vesicular stomatitis virus G-protein (VSV-G) enhances transduction efficiency, optimal expression strategies during the Autographa californica multiple nucleopolyhedrovirus (AcMNPV) infection cycle remain unexplored. This study investigates how VSV-G expression timing affects pseudotype incorporation into budded virions (BVs) and subsequent transduction efficacy. Methods: Three recombinant AcMNPV constructs were generated, each expressing VSV-G under distinct baculoviral promoters (ie1, gp64, and p10) and GFP via a CMV promoter. VSV-G incorporation was verified by Western blot, while transduction efficiency was quantified in mammalian cell lines (fluorescence microscopy/flow cytometry) and rat hind limbs. Viral productivity was assessed through production kinetics and plaque assays. Results: All the pseudotyped viruses showed significantly enhanced transduction capacity versus controls, strongly correlating with VSV-G incorporation levels. The p10 promoter drove the highest VSV-G expression and transduction efficiency. Crucially, BV production yields and infectivity remained unaffected by VSV-G expression timing. The in vivo results mirrored the cell culture findings, with p10-driven constructs showing greater GFP expression at low doses (10⁴ virions). Conclusions: Strategic VSV-G expression via very late promoters (particularly p10) maximizes baculoviral transduction without compromising production yields. This study establishes a framework for optimizing pseudotyped BV systems, demonstrating that late-phase glycoprotein expression balances high mammalian transduction with preserved insect-cell productivity—a critical advancement for vaccine vector development. Full article

Objectives: The anticancer effects of PI3Kα inhibitors (PI3Ki) are constrained by their hyperglycemic side effects, while the efficacy of conventional hypoglycemic agents, such as insulin, metformin, and SGLT-2 inhibitors, in mitigating PI3Ki-induced hyperglycemia remains suboptimal. Dorzagliatin, a novel glucokinase activator, has been approved in China for the management of hyperglycemia, offering a promising alternative. This study aims to investigate the pharmacokinetic properties and potential mechanisms of drug interactions of dorzagliatin in the regulation of PI3K-induced hyperglycemia. Methods: Plasma concentrations of WX390, BYL719, and Dorz in mice were measured using high performance liquid chromatography-tandem mass spectrometry (LC-MS/MS) analysis. Pharmacokinetic (PK) parameters and PK/PD models were derived by using Phoenix WinNonlin 8.3.5 software. Blood glucose levels at various time points and tumor volume changes over a four-week period were assessed to explore the interactions when PI3Ki were combined with dorzagliatin. Results: The results indicated that, compared to the Dorz group, the combination groups (Dorz + BYL719, Dorz + WX390) exhibited increases in ${\mathrm{A}\mathrm{U}\mathrm{C}}_{0\to t}$ of dorzagliatin by 41.65% and 20.25%, and in C_max by 33.48% and 13.32%, respectively. In contrast, co-administration of these PI3Ki with dorzagliatin resulted in minimal increase in their plasma exposure. The combination therapy group (Dorz+BYL719) exhibited superior antitumor efficacy compared to the BYL719 group. Conclusions: Our findings indicate that the drug–drug interactions (DDIs) between dorzagliatin and multiple PI3Ki (including WX390 and BYL719) may partially account for the enhanced antitumor efficacy observed in the combination therapy group compared to PI3Ki monotherapy. This interaction may be explained by the inhibition of P-glycoprotein (P-gp) and the pharmacological mechanism of dorzagliatin regarding the activation of insulin regulation. Full article

The Baculovirus Expression Vector System (BEVS) is an important protein and complex biologics production platform. The baculovirus GP64 protein is the major envelope glycoprotein that aids in virus entry and is required for cell-to-cell transmission in cell culture. Several studies have developed strategies around gp64 gene disruption in an attempt to minimize baculovirus co-production. Here, we investigate the result of transiently targeting the baculovirus gp64 gene with CRISPR-Cas9 during infection. Because not all genomes are effectively disrupted, we describe a variant calling methodology that allows the detection of the targeted mutations in gp64 even though these mutations are not the dominant sequences. Using a transfection-infection assay (T-I assay), the AcMNPV gp64 gene was targeted at six different locations to evaluate the effects of single and multiple targeting sites, and we demonstrated a reduction in the levels of baculovirus vectors while maintaining or enhancing foreign protein production when protein was driven by a p6.9 promoter. Viral genomes were subsequently isolated from the supernatant and cell pellet fractions, and our sequencing pipeline successfully detected indel mutations within gp64 for most of the single-guide RNA (sgRNA) targets. We also observed that 68.8% of variants found in the virus stock were conserved upon virus propagation in cell culture, thus indicating that they are not detrimental to viral fitness. This work provides a comprehensive assessment of CRISPR-Cas9 genome editing of baculovirus vectors, with potential applications in enhancing the efficiency of the BEVS. Full article

Multidrug resistance (MDR) poses a significant challenge in cancer therapy, often leading to treatment failure and relapse. ATP-binding cassette (ABC) transporters, particularly ABCG2, play a pivotal role in MDR development by actively expelling chemotherapeutic agents from cancer cells. This study investigates the effects of two groups of primaquine derivatives—fumardiamides (1a–d) and bis-ureas (2a, b), both bearing halogenated benzene rings—on the activity of P-glycoprotein (P-gp) and ABCG2. Their potential to reverse MDR was evaluated through a series of functional assays aimed at comparing transporter–compound interactions. The results indicated that fumardiamide derivatives, specifically 1a, 1b, and 1d, exhibited potent inhibition of ABCG2 while having no effect on P-gp, demonstrating a selective mode of action. The tested derivatives displayed low to moderate cytotoxicity and did not affect ABCG2 expression or localization. Moreover, these compounds enhanced the sensitivity of drug-resistant cancer cell lines to mitoxantrone, underscoring their potential to overcome ABCG2-mediated MDR. These findings suggest that chemical modifications of primaquine, particularly the incorporation of fumardiamide moieties, confer novel biological properties, providing promising leads for the development of selective ABCG2 inhibitors. Full article

In recent decades, the keeping of exotic animals has gained popularity among enthusiasts worldwide. However, alongside the development of exotic animal husbandry, issues related to health status and adequate veterinary care are coming to the forefront. The introduction of new snakes into a collection and shared enclosures should always be preceded by an assessment of their parasitic status. In our study, we present an overview of the screening for the presence of Cryptosporidium spp. in individuals captured in regions of Indonesia and Suriname, intended for further trade. Out of 40 tested fecal samples, the presence of cryptosporidial oocysts was confirmed in 6 samples. Detection was performed by molecular methods, namely Nested PCR targeting the GP60 gene region (60 kDa glycoprotein). By sequencing, we confirmed the presence of C. parvum in Oligodon octolineatus (n = 1) and Trimeresurus insularis (n = 1), C. tyzzeri in Corallus spp. (n = 2), and C. hominis in Boiga dendrophila spp. gemmicincta (n = 2), which is the very first time that this species has been detected in snakes in captivity. Although the presence of Cryptosporidium species, typical for snakes, was not detected, the identified species may pose a health risk to humans, especially workers who come into direct contact with animals. Full article

Porcine reproductive and respiratory syndrome virus (PRRSV) modified live virus (MLV) vaccination is used to control PRRSV. In China, farms conduct random sampling from sow herds every 4 to 6 months. They use the enzyme-linked immunosorbent assay (ELISA) method to monitor the immune status of the herd by tracking the positive rate or the sample-to-positive ratio. However, in farms that implement mass vaccination and have stable production, the positive rate of ELISA antibodies has decreased, especially in high-parity sows. This poses a considerable challenge to the current monitoring approach of PRRSV immunity. It remains unclear whether this reflects insufficient sensitivity of the kits for these special scenarios or the fact that the sows have truly lost immunity. In this study, 233 samples from four farms (A–D) across different regions of China were acquired. They were tested using four representative ELISA kits, two targeting the nucleocapsid protein (N) and two targeting the glycoprotein (GP) to evaluate PRRS immune status. The respective sample positive rates in A–D were 57.1–100%, 50.9–100%, 50–100%, and 75.7–100% using the kits. The positive rates using the four ELISA kits were 50.0–75.7%, 70.0–75.7%, 82.5–97.1%, and 100%, respectively, with poor agreement among them. The positive rates and humoral antibody levels for parity 1 and 2 sows were significantly lower than those with higher parities (>4). Eighty-eight ELISA-negative samples identified using ELISA kit A were verified using a viral neutralizing test (VNT), with only 15.9% of the samples testing negative. In conclusion, the ELISA antibody negativity issue existed, mostly occurring in specific farms tested using a specific kit. However, the low correlation with the VNT results and the poor agreements among the kits suggest that relying on one ELISA test is insufficient to monitor the immune status of PRRSV MLV-vaccinated herds. Full article

P-glycoprotein (P-gp), a transmembrane efflux pump encoded by the ABCB1/MDR1 gene, is a major contributor to multidrug resistance in hematological malignancies. These malignancies, arising from hematopoietic precursors at various differentiation stages, can manifest in the bone marrow, circulate in the bloodstream, or infiltrate tissues. P-gp overexpression in malignant cells reduces the efficacy of chemotherapeutic agents by actively expelling them, decreasing intracellular drug concentrations, and promoting multidrug resistance, a significant obstacle to successful treatment. This review examines recent advances in combating P-gp-mediated resistance, including the development of novel P-gp inhibitors, innovative drug delivery systems (e.g., nanoparticle-based delivery), and strategies to modulate P-gp expression or activity. These modulation strategies encompass targeting relevant signaling pathways (e.g., NF-κB, PI3K/Akt) and exploring drug repurposing. While progress has been made, overcoming P-gp-mediated resistance remains crucial for improving patient outcomes. Future research directions should prioritize the development of potent, selective, and safe P-gp inhibitors with minimal off-target effects, alongside exploring synergistic combination therapies with existing chemotherapeutics or novel agents to effectively circumvent multidrug resistance in hematological malignancies. Full article