Pakistan Journal of Pharmaceutical Sciences

Abstract: Background: Epithelial-mesenchymal transition (EMT) is involved in colorectal cancer (CRC) pathogenesis. Curcumin (CUR) is widely used in the treatment of CRC. Polystyrene nanoparticles (PS-NPS) have a high specific affinity and have an inhibitory effect on cell proliferation. Therefore, the purpose of this study is to investigate the mechanism of inhibition of EMT in colorectal cancer mice by preparing curcumin polystyrene nanoparticles (CUR-PS-NPS). Objective: This study aimed to construct CUR-PS-NPS and investigate their mechanism in inhibiting CRC EMT via the miR-186/PI3K/AKT/Twist axis. Method: CUR-PS-NPS were prepared and characterized. In-vivo, CRC model mice (Balb/c, male) were divided into four groups: sham-operated control (PC), CRC model (CRC), low-dose CUR-PS-NPS (Cur-PS-NPS-L), and high-dose CUR-PS-NPS (Cur-PS-NPS-H). After intraperitoneal injection, tumor volume, EMT marker proteins and changes in miR-186 and the PI3K/AKT/Twist pathway were measured. In-vitro, human normal colonic epithelial cells (NCM460) were used to validate miR-186 interference and SW480 cells were co-transfected with a PI3K inhibitor to clarify the specific mechanism by which CUR-PS-NPS inhibit EMT. Results: In CRC mice intervened with CUR-PS-NPS, the tumor volume was reduced by 50%, the expression of mir-186 was up-regulated by 2.5 times, the phosphorylation level of PI3K/AKT was decreased by 40%, the expression of  Twist protein was down-regulated by 35% and the expression of the EMT marker E-cadherin was increased. The expressions of N-cadherin, Vimentin and Snail decreased significantly. Conclusion: CUR-PS-NPS can effectively delay the EMT process of CRC by up-regulating mir-186, inhibiting the PI3K/AKT signaling pathway and the expression of Twist. This discovery provides a new strategy for targeted therapy based on nanocarriers and its application potential in clinical practice can be further explored in the future.

Abstract: Background: Fine particulate matter (PM2.5) induces lung inflammation through pathways involving receptor for advanced glycation end-products (RAGE), S100A9 and nuclear factor-kappa B (NF-κB). Xing Su San and Sang Xing Tang are traditional Chinese medicine formulas used for respiratory disorders associated with cool-dry and warm-dry conditions, respectively. Objectives: To investigate whether treatment with these formulas is associated with reduced expression of RAGE, S100A9 and NF-κB in lung tissue of rats exposed to PM2.5 under simulated warm-dry and cool-dry conditions. Methods: Forty male Wistar rats were divided into five groups (n=8 each): blank control, warm-dry model, cool-dry model, Sang Xing Tang treatment and Xing Su San treatment. Exposed groups received PM2.5 (500 µg/m³, 3 hours/day, 28 days) under warm-dry (20°C, 33% humidity) or cool-dry (8°C, 33% humidity) conditions. Treatment groups received oral gavage of Sang Xing Tang (8.5 g/kg) or Xing Su San (12.5 g/kg) before exposure. RAGE messenger RNA was measured by reverse transcription quantitative polymerase chain reaction, NF-κB p65 protein by Western blot and S100A9 by immunohistochemistry. Results: Both PM2.5 exposure conditions were associated with significantly increased RAGE (2.8-fold and 2.5-fold), NF-κB p65 (2.4-fold and 2.2-fold) and S100A9-positive cells (3.45% and 3.13%) compared to controls (0.82%; all P < 0.05). Sang Xing Tang treatment was associated with reduced markers in the warm-dry model (RAGE: 1.6-fold; NF-κB: 1.5-fold; S100A9: 2.25%). Xing Su San treatment was associated with reduced markers in the cool-dry model (RAGE: 1.5-fold; NF-κB: 1.4-fold; S100A9: 2.08%; all P < 0.05). Conclusion: Xing Su San and Sang Xing Tang attenuate PM2.5-induced upregulation of RAGE, S100A9 and NF-κB in rat lung tissue in a pattern-specific manner. These hypothesis-generating findings provide preliminary evidence warranting further mechanistic validation.

Abstract: Background: Ferroptosis-related oxidative injury has been implicated in the progression of heart failure (HF). Rosmarinic acid (RA) has antioxidant and anti-lipid peroxidation properties, but its role in HF-associated ferroptosis-related cardiomyocyte injury remains unclear. Objectives: This study aimed to investigate whether RA attenuates Ang II-induced cardiomyocyte injury by modulating ferroptosis-related oxidative injury and GPX4/xCT-associated defense responses. Methods: RA-related targets were retrieved from TCMSP, while HF-related and ferroptosis-related genes were obtained from GeneCards/OMIM and FerrDB. Intersection targets were analyzed using PPI network construction, GO and KEGG enrichment analyses, RA-target-disease-pathway network construction and molecular docking. An Ang II-induced H9c2 cardiomyocyte injury model was established for validation. Cell viability was assessed by CCK-8 assay; ANP and BNP mRNA levels were measured by RT-qPCR; MDA content was measured to evaluate lipid peroxidation; and GPX4 and xCT protein expression was examined by Western blotting. Results: Twenty-six RA-related targets, 3311 HF-related genes and 1393 ferroptosis-related genes were identified. Intersections yielded 397 HF-associated ferroptosis targets and 9 RA-related candidate targets. Network analysis highlighted PPARG, PTGS2, MAPK1, CDKN1A and CASP3 as important targets. Enrichment analyses indicated associations with oxidative stress, apoptotic regulation, iron ion response, inflammatory signaling, lipid and atherosclerosis, TNF, IL-17 and AGE-RAGE pathways. Molecular docking suggested favorable binding of RA to these targets, especially PPARG and PTGS2. In Ang II-treated H9c2 cells, RA restored cell viability, reduced ANP and BNP expression, decreased MDA accumulation and increased GPX4 and xCT expression; these effects were partially reversed by erastin. Conclusion: RA may attenuate Ang II-induced cardiomyocyte injury by modulating ferroptosis-related oxidative injury and GPX4/xCT-associated defense responses. These findings provide preliminary computational and cellular evidence supporting RA as a candidate compound for further investigation in HF-related cardiomyocyte injury.

Abstract: Background: Exosomes derived from bone marrow mesenchymal stem cells (BMSCs) can deliver microRNAs to recipient cells and modulate their functions; however, investigations into their molecular effects have frequently relied on surrogate cancer cell lines, whose relevance to benign CAG pathology remains uncertain and must be explicitly acknowledged. Objectives: To explore whether BMSC-derived exosomal miR-618-3p influences proliferation, apoptosis and epithelial-mesenchymal transition (EMT). Methods: Twelve CAG tissues and six normal gastric mucosa samples were obtained from the laboratory tissue bank. The human gastric adenocarcinoma cell line SGC-7901 was used as an in-vitro malignant surrogate for exploratory mechanistic evaluation, not as a model of benign CAG. BMSC-derived exosomes (BMSC-exo) were isolated by ultracentrifugation and characterized. CAG cells were treated with control exosomes or exosomes enriched with miR-618-3p mimic. miR-618-3p and TFF3 expression were detected by RT-qPCR and Western blot. Cell proliferation was assessed by CCK-8, apoptosis by caspase-3 activity and epithelial-mesenchymal transition (EMT) markers by RT-qPCR. A dual-luciferase reporter assay validated the interaction between miR-618-3p and the TFF3 3′UTR. Results: miR-618-3p was decreased and TFF3 increased in CAG tissues (both p < 0.05). The dual-luciferase assay suggested a direct interaction between miR-618-3p and the TFF3 3′UTR. Unexpectedly, unmodified BMSC-exo treatment reduced miR-618-3p expression in recipient SGC-7901 cells, increased TFF3 protein levels, enhanced proliferation, reduced caspase-3 activity, decreased E-cadherin and increased vimentin expression (all p < 0.05. In contrast, BMSC-exo enriched with miR-618-3p mimic increased miR-618-3p levels, suppressed TFF3, reduced proliferation, increased caspase-3 activity, upregulated E-cadherin and downregulated vimentin (all p < 0.05). Conclusion: In this preliminary in-vitro study using a gastric adenocarcinoma cell line, BMSC-exo miR-618-3p was associated with reduced TFF3 expression and altered proliferation, apoptosis and EMT markers. These findings describe a potential regulatory axis but remain strictly exploratory.

Abstract: Background: Moderate hamstring strain is a common sports injury impairing daily and athletic function. While topical nonsteroidal anti-inflammatory drugs (NSAIDs) offer localized pain relief with reduced systemic effects, high-quality evidence specific to hamstring injury is limited. Objectives: To evaluate the effect of a short-course topical diclofenac epolamine patch combined with rehabilitation on pain and functional recovery in moderate hamstring strain. Methods: In this single-center, double-blind, placebo-controlled trial, 120 patients were randomized to an experimental group (diclofenac patch twice daily for 14 days plus rehabilitation) or a control group (placebo patch plus identical rehabilitation). Outcomes assessed included pain measured by the Visual Analogue Scale (VAS), as the primary endpoint, muscle strength, knee range of motion (ROM), functional back-kick test, time to rehabilitation criteria and safety (adverse events, liver/renal function). Results: The diclofenac group demonstrated significantly greater pain reduction at days 14, 21 and 28 post-intervention (all p < 0.01). Muscle strength recovery was superior at days 21 and 28, knee ROM improved more at day 28 and functional back-kick performance was better at day 28 (all p < 0.05). Time to meet rehabilitation criteria was significantly shorter in the experimental group (p = 0.002). Adverse event incidence and changes in liver/renal function were similar between groups (all p > 0.05). Conclusion: A short-course topical diclofenac epolamine patch, combined with rehabilitation training, effectively alleviates pain, accelerates functional recovery, and is safe for treating moderate hamstring strain, supporting its use in clinical and sports rehabilitation.