Pakistan Journal of Pharmaceutical Sciences

Abstract: Background: Dysregulated expression of prolyl 4-hydroxylase subunit alpha 3 (P4HA3) has been identified in Colon cancer, breast cancer, head and neck cancers, etc. and is closely associated with tumor progression. However, its specific roles in gastric cancer (GC), particularly regarding alterations in metabolic pathways, are remain unknown. Objectives: This study investigated how P4HA3 influences glycolytic metabolic reprogramming and the proliferation and invasion of gastric cancer cells via the PI3K/AKT/mTOR signaling pathway. Methods: To determine P4HA3 levels in GC tissues and corresponding cell lines, bioinformatics methods, immunohistochemical (IHC) staining and Western blotting (WB) were conducted. Furthermore, gain- and loss-of-function experiments, including cell viability assays (CCK-8), colony-forming tests, scratch wound assays and Transwell invasion experiments, were utilized to explore the influence of P4HA3 on GC cell growth, motility and invasive capacity. Changes in cellular metabolism were assessed using the Seahorse system (under normoxic conditions), including measurements of basal metabolism, glycolytic capacity and mitochondrial respiration (oxidative phosphorylation), as well as Western blot analysis of key glycolytic proteins. Additionally, the involvement of the PI3K/AKT/mTOR signaling axis was examined by gene set enrichment analysis (GSEA), WB analysis and pharmacological inhibition studies. Results: GC tissues and cell lines exhibited markedly increased levels of P4HA3, correlating significantly with decreased overall survival (OS). Experimental data indicated that P4HA3 facilitated GC cell proliferation, migration and invasion. In addition, P4HA3 enhanced glycolytic activity and increased the expression of key glycolysis-related proteins. Unexpectedly, P4HA3 also increased mitochondrial respiration under the assay conditions, suggesting that P4HA3 may induce a broader metabolic activation rather than an exclusively glycolytic shift. Mechanistically, the glycolysis-enhancing effects of P4HA3 were found to require PI3K/AKT/mTOR signaling (as shown by inhibitor studies). Conclusion: P4HA3 promotes glycolysis via the PI3K/AKT/mTOR signaling pathway, thereby promoting the progression of gastric cancer.

Abstract: Background: Bacteria have become more resistant to commonly used antibiotics. Among the many reasons, biofilm-mediated protection plays an indispensable role. To overcome microbial resistance, alternative antibiofilm therapies may be developed to address this challenge and they should be cost-effective and readily available. Objectives: To evaluate the antibiofilm activities of natural compounds Allium Cepa (Onion), Menthe spicata (mint) and Zingiber officinale (Ginger) against Escherichia coli and Pseudomonas Aeruginosa Biofilms. Methods: This cross-sectional study includes 150 patients from various clinics in Karachi, Pakistan. Samples of Escherichia coli and Pseudomonas aeruginosa were collected and the study was carried out to determine the antibiofilm-forming activity of natural product extracts of Allium cepa (Onion) G.H.NO 100929, Mentha spicata (mint) G.H.NO100933 and Zingiber officinale (Ginger) G.H.NO100928, evaluated separately and in different combinations. 96 well plate method was used to evaluate biofilm forming activity of microorganisms and antibiofilm forming activity of Allium Cepa (Onion) Menthe Spicata( mint) Zingiber Officinale (Ginger Peels) Results: Natural compounds Allium Cepa (Onion) Menthe Spicata( mint) Zingiber Officinale (Ginger) has shown antibiofilm activity against Escherichia coli and Pseudomonas aeruginosa biofilms separately but when used in combination only synergistic activity shows ginger and mint combination with FIC 0.45 against Pseudomonas Aeruginosa. Whereas both organisms have shown resistance to conventional antimicrobial agents. Conclusion: This study concludes that Allium Cepa (Onion) Menthe Spicata (mint) Zingiber Officinale (Ginger) having antibiofilm activity when used separately but in combination Zingiber Officinale (Ginger) And Menthe Spicata (mint) against Pseudomonas Aeruginosa only shows synergistic activity where as other combination shows indifferent activity but shows lower concentration of extract as compare to when used separately but not match the criteria of synergism but importantly none of combination shows antagonist activity. These findings emphasize the need to incorporate these natural agents, after drug authority approval, with already available antimicrobial agents.

Abstract: Background: Suaeda fruticosa is a widely distributed plant in Pakistan which has been traditionally used as a remedy for constipation, dyspnoea, headache, nausea, neurosis and vertigo. Objectives: The present study aimed to scientifically validate the traditionally claimed medicinal properties of Suaeda fruticosa. Methods: The experiment involved the preparation of a 70% ethanolic extract of Suaeda fruticosa (Sf.Cr), which was tested ex-vivo in an isolated rabbit jejunum preparation for spasmolytic activity. Moreover, the crude extract was tested on isolated tracheal and aorta preparations to evaluate the effects. Results: Sf.Cr dose-dependently relaxed the jejunum and inhibited K⁺(80 mM)-induced contractions with EC₅₀ value of 0.229 mg/ml and 0.095 mg/ml, respectively. Further, the Sf.Cr caused a rightward shift in Ca2+ concentration response curves (CRCs), similar to verapamil. So, calcium channel-blocking and anticholinergic actions were responsible for the antispasmodic (anti-diarrhoeal) effect. Sf.Cr produced inhibition of carbachol (1 µM)- and K⁺(80 mM)-induced contractions in isolated rabbit tracheal tissues with EC₅₀ values of 0.8339 mg/ml and 1.022 mg/ml, respectively. Sf. Cr also completely abolished the Phenylephrine (1 µM)- and K+(80 mM)-induced constrictions in isolated rabbit aortic rings, with EC50 values of 0.9448 mg/ml and 0.2026 mg/ml, respectively. Molecular docking against calmodulin (PDB ID: 6M7H) showed that 3-hydroxy-2-quinolinone attained the highest glide score (-6.742 kcal.mol⁻¹), establishing hydrogen bonding and Pi–Pi stacking interactions with MET124 and PHE92, surpassing the binding affinity of the standard drug verapamil (-5.464 kcal.mol⁻¹). SwissADME profiling showed that all identified phytoconstituents complied with Lipinski’s rule of five, exhibited high gastrointestinal absorption and were predicted to cross the blood-brain barrier, supporting their oral bioavailability as drug candidates. Conclusion: These findings collectively established Sf.Cr as a source of non-specific calcium channel-blocking phytochemicals with potential for developing antispasmodic, bronchodilator, and vasorelaxant agents.

Abstract: Background: Ulcerative colitis is a relapsing inflammatory bowel disease driven by dysregulated immunity, epithelial barrier injury and microbial imbalance. Current therapies remain limited by relapse, adverse effects and incomplete response. Quercetin has anti-inflammatory and barrier-protective activities and ERK-MMP signaling may link inflammation to mucosal tissue damage. Objectives: To evaluate whether the protective effects of quercetin in experimental ulcerative colitis (UC) are associated with modulation of ERK-MMP signaling, while avoiding causal overinterpretation beyond the animal and cellular evidence. Methods: A dextran sulfate sodium (DSS)-induced mouse model and a lipopolysaccharide (LPS)-stimulated RAW264.7 macrophage inflammatory model were used. Mice were allocated to the control, UC model and low-, medium- and high-dose quercetin groups. Disease activity index (DAI; unitless), colon length, histopathology, inflammatory cytokines and ERK/MMP-related molecules were evaluated by histological staining, ELISA, qRT-PCR and western blotting. In-vitro, quercetin was compared with the ERK phosphorylation inhibitor U0126 and an agonistic stimulus to explore pathway-related associations. The study was exploratory and did not include genetic rescue experiments or human validation. Results: In DSS-treated mice, medium- and high-dose quercetin were associated with improved colon morphology, reduced DAI scores and partial restoration of colon length. Quercetin decreased pro-inflammatory cytokine expression and increased IL-10 expression. In both mouse colon tissues and LPS-stimulated RAW264.7 cells, quercetin was associated with reduced ERK1/2 activation and lower MMP-3 and MMP-9 expression, together with partial recovery of TIMP expression. These molecular changes were directionally consistent with the effects observed after ERK inhibition in-vitro. Conclusion: Quercetin showed protective effects in experimental UC models and these effects may be associated with modulation of the ERK-MMP signaling axis. Further pathway-specific rescue experiments, pharmacokinetic studies and human-relevant models are required before this mechanism can be considered definitive or clinically translatable.

Abstract: Background: Laxatives are the mainstay for pediatric functional constipation (FC), yet response factors need more real-world research. Objectives: This study investigated factors associated with laxative treatment response in children with functional constipation. Methods: Retrospective data were screened from 103 pediatric FC patients receiving polyethylene glycol treatment (The First Hospital of Qinhuangdao, Feb 2023–Nov 2025). After exclusion, 100 cases were included and divided into the responder group (n=68) and the non-responder group (n=32). The primary outcome measures included the Pediatric Constipation Score (PCS), the Bristol Fecal Morphology Scale (BSFS) and defecation frequency. Secondary endpoints included the incidence of abdominal pain, incidence of bloating, average daily dietary fiber intake, average daily water intake and average daily physical activity. A logistic regression analysis was conducted to identify independent risk factors associated with treatment response. Results: The baseline clinical characteristics of both groups were well-balanced (all P > 0.05). Post-treatment evaluation showed that the PCS score in the responder group decreased more significantly compared to the non-responder group (P<0.001). Meanwhile, increases in bowel movement frequency and BSFS scores were more pronounced in the responder cohort (all P<0.001). Compared to the non-responder group, the responder group also had lower incidences of abdominal pain and distension (all P<0.05). In terms of lifestyle-related indicators, the responder group had significantly higher daily average dietary fiber intake, average daily water intake and average daily physical activity levels (all P<0.001). Results from logistic regression analysis indicated that high baseline PCS scores, elevated incidence of abdominal pain, insufficient dietary fiber intake, low daily water intake and inadequate physical activity emerged as independent risk factors for poor laxative response in pediatric FC patients (all P<0.05). Conclusion: Laxative response in pediatric FC is affected by several factors, such as severe constipation, poor diet, low activity and abdominal pain.