Cell and Tissue Journal

Abstract Aim: Oregano (Mentha longifolia) is an important medicinal plant from the mint family, which is widely used in food and pharmaceutical industries due to its active compounds. As an efficient approach, the use of biological elicitors (yeast, bacterial, and fungal agents) as well as non-biological elicitors (inactive enzymes, ultraviolet rays, heavy metal salts, polysaccharides, salicylic acid, jasmonic acid, and melatonin) can provide a way to increase the production of secondary metabolites in plant cell cultures. Given the importance of Oregano in the pharmaceutical and therapeutic industries, this study was carried out to assess melatonin's effect on the percentage of cell viability and the level of important bioactive compounds of oregano.  
Materials and methods: This study was performed in a factorial experiment with a completely randomized design and three replications in the lab. Melatonin (0, 100, and 200 μM based on the previous studies) was added to the MS culture medium containing 2,4-D (1 mg.L^-1). Then, 24, 48, and 72 h after treatment, the percentage of cell survival and the level of important oregano compounds were measured by tetrazolium and HPLC tests, respectively. In SAS software version 9.4, analysis of variance (ANOVA) was used to analyze the data and the LSD method to compare the means.
Results: Based on the results of the tetrazolium test, there was no significant difference in the percentage of cell viability in various concentrations of melatonin. However, the survival percentage of cells decreased with the passage of time, so that the lowest survival percentage was observed 72 h after melatonin treatment. Adding melatonin to the culture medium increased the production of secondary metabolites. At the concentration of 100 µM, the amount of menthol, menthone, pulegone, and 1,8-cineole increased by 77, 108, 46, and 80%, respectively, after 48 hours in contrast to the control. The increase in the level of menthol, menthone, pulegone, and 1,8-cineole compounds in the concentration of 200 µM of melatonin was respectively 125, 130, 140, and 120%, after 48 hours when compared to the control. Therefore, increasing melatonin concentration from 100 to 200 µM in cell culture significantly increased the level of compounds compared to the control. The levels of menthol, menthone, pulegone, and 1,8-cineole increased up to 48 hours after treatment, but after that time, they showed a significant decrease. Since the ultimate goal is to increase the production of biologically active compounds and not the highest cell viability, therefore, 48 hours after applying the treatment of 200 μM melatonin provides the best conditions for obtaining a high level of valuable oregano metabolites. The above treatment increased menthol, menthone, pulegone, and 1,8-cineole by 125, 130, 140, and 120%, respectively.
Conclusion: In this study, for the first time, the effect of melatonin elicitor on important secondary metabolites of the Oregano plant was investigated to determine the potential of using this metabolic stimulant in research and commercial areas. Overall, The use of melatonin at a concentration of 200 μM along with the extraction of metabolites 48 hours after the treatment can enhance menthol, menthone, pulegone, and 1,8-cineol, while maintaining the viability of oregano cells

Abstract Aim: Cannabis sativa L. contains valuable cannabinoids necessary for industrial, nutraceutical  and pharmaceutical applications. To ensure the availability of cannabis plant materials in vitro culture approaches guarantee rapid and mass production to meet the growing demand. On the other hand, callogenesis can also guarantee the production of secondary metabolites in vitro through suspension cell culture. The goal of the current study was to compare different basal culture media and plant growth regulator combinations to offer the best conditions for inducing callus formation in medical cannabis.
Material and Methods: Leaf explants were placed on MS and DKW basal culture media were used along with B5 vitamins, 30 g/l sucrose, and 7 g/l of Agar, together with various concentrations of plant growth regulators, including 2,4-D (2, 5, and 8 µmol/l) and BA (0.5 and 2.5 µmol/l).
Results: The calli appeared between 5 to 7 days after establishment on various basal media supplemented with different types and concentrations of hormones. The presence of plant growth regulators is essential for callus formation in this plant, as the cultured explants in control treatments without hormones did not produce any calli. Callus formation was observed in all media and hormonal treatments, but produced calli were different from each other in terms of size, color and texture. Our study showed that texture of calli was compact or friable. The texture of calli in DKW was the most compact with the least amount of cytokinin at any concentration of auxin. However, in MS, it was the most friable with the highest amount of cytokinin at any concentration of auxin. Also, in this medium, the texture of calli was the most friable with the highest concentration of auxin. The color of calli in hormonal treatments present in MS culture medium was white and light cream, but in most hormonal treatments in DKW culture medium, it was white. The results of this study indicate that the best medium was MS medium supplemented with 2 (µmol/l) 2,4-D with 2.5 (µmol/l) BA (friable callus). Furthermore, the most inappropriate medium for callus formation from cannabis leaves in terms of all the characteristics examined was the DKW culture medium along with 5 (µmol/l) 2,4-D in combination with 2.5 (µmol/l) BA. The compression of calli can be caused by the reduction of proliferation in the cells that are dividing, this reaction can be influenced by the auxin inside the explant. Adding high auxin to endogenous auxin, in addition to adding cytokinin with a low concentration can affect the formation of compact texture. The formation of friable callus requires a balanced combination of auxin and cytokinin. Another important factor in plant tissue culture is the color of the calli, which is a sign of tissue health and vitality. For example, a healthy calli under light conditions is usually green in color, but a dark brown or black color is usually a sign of the death of the calli, which is due to pollution, stress or the production of substances such as phenols. Maximizing callus formation in the shortest possible time is one of the important objectives in tissue culture techniques, which not only saves time and cost but also prevents possible somaclonal variations. In general, according to the results, one of the suitable mediums for callus production in cannabis is the DKW,so the results of this study confirm the introduction of  DKW as the best culture medium in past studies. However, the MS culture medium is also the best medium in the available study and the least suitable result of this study is related to one of the hormonal combinations in the DKW culture medium. This difference with the mentioned study may be due to differences in genotype, laboratory materials, hormone amount, vitamins or other conditions affecting the tissue culture of this plant.
Conclusion: Calli provides the opportunity to reproduce plants on a large scale and produce disease-free plants. Generally according to the plant, friable calli have a more suitable tissue for somatic embryogenesis and suspension culture, whereas compact calli have a higher potential for regeneration. This medium is recommended for future studies on somatic embryogenesis and cell suspension cultures due to the calli being friable in it. 

Abstract Introduction: Paxlovid, an investigational oral therapeutic combining the antiviral Nirmatrelvir and the pharmacokinetic enhancer Ritonavir, is being developed to treat SARS-CoV-2 infection, aiming to reduce severe disease progression, hospitalization, and mortality. While its efficacy has been shown in Phase 2/3 trials for non-hospitalized patients, its safety, particularly regarding fetal development, remains under investigation. Coronaviruses have long posed significant challenges to global public health, with their potential to cause severe respiratory infections in humans. Over the past two decades, two novel coronaviruses—Severe Acute Respiratory Syndrome Coronavirus (SARS-CoV) and Middle East Respiratory Syndrome Coronavirus (MERS-CoV)—have emerged, leading to widespread morbidity and mortality. The SARS - CoV outbreak, which occurred in the early 2000s, infected over 8,000 individuals worldwide and resulted in nearly 800 deaths, representing a mortality rate of approximately 10 %. Similarly, MERS - CoV, identified in 2012, caused 857 confirmed infections and 3 34 deaths, with a strikingly high mortality rate of around 35 %. These outbreaks underscored the persistent threat posed by coronaviruses to human health.
Aims: Pregnant women represent a particularly vulnerable population during infectious disease outbreaks, and the potential teratogenic effects of new therapeutics must be carefully evaluated. Previous studies on Nirmatrelvir have explored its effects on skeletal morphogenesis, but comprehensive data on Paxlovid's impact on fetal development remain limited. This study aims to address this gap by investigating the effects of Paxlovid on the reproduction and skeletal morphogenesis of rat embryos, providing critical insights into its safety profile during pregnancy. By elucidating the potential risks associated with Paxlovid, this research contributes to the broader effort to ensure the safe use of antiviral therapies in vulnerable populations. This study assessed the potential embryotoxic effects of Paxlovid on rat fetal skeletal morphogenesis, following ICH guidelines.
Materials and methods: Pregnant rats were divided into four groups, receiving doses of 0, 100/60, 300/200, or 1000 mg/kg/day of Paxlovid. Maternal clinical symptoms and weight were monitored, and fetal outcomes, including weight, crown-rump length (CRL), and abdominal circumference (AC), were evaluated on gestational day 21. Histological analysis of fetal skeletal tissue was conducted using hematoxylin-eosin and Alizarin Red S staining to detect structural abnormalities.
Results: Results showed reduced maternal weight gain in Paxlovid-treated groups compared to controls. Fetuses in treated groups also exhibited lower weight, CRL, and AC. However, histological analysis revealed no structural abnormalities in skeletal tissue. These findings suggest that while Paxlovid may transiently affect fetal growth metrics, it does not induce teratogenic effects on skeletal development.
Discussion: The findings of this study revealed no histological abnormalities in the skeletal system of fetuses exposed to Paxlovid. The observed effects were limited to reductions in fetal weight, crown-rump length (CRL), and abdominal circumference (AC). Collectively, the data from this investigation, along with existing evidence, suggest a low risk of fetal harm associated with Paxlovid, which comprises Nirmatrelvir (NMV), a potent and selective inhibitor of the SARS-CoV-2 main protease, and Ritonavir, a pharmacokinetic enhancer. Nonetheless, the use of Paxlovid during pregnancy should be carefully evaluated, with a thorough discussion of potential risks and benefits conducted in consultation with a healthcare professional.
Conclusion: The study concludes that Paxlovid does not compromise the histological integrity of fetal skeletal tissue, supporting its potential safety during pregnancy. However, further research is needed to understand the mechanisms behind the observed growth effects and to confirm these findings in human populations. Overall, the results indicate a low risk of fetal harm, reinforcing Paxlovid's safety profile for use in pregnant individuals

Abstract Aims: Salvia L. with more than 1000 taxa is the largest genus in Lamiaceae family, that naturally grows in diverse regions of the world. The genus is represented in Iran by about 60 species, which 17 of them are endemic for the country. Salvia macrosiphon Boiss., is an annual aromatic herb of this genus, which is widely distributed in diverse parts of Iran. Due to the presence of a wide range of secondary metabolites (especially essential oil), this plant has been widely applied in the traditional medicine to cure different diseases. Various types of glandular and non-glandular trichomes have been detected in Lamiaceae taxa.  Essential oils are composed of several compounds, and are biosynthesized  and stored in the glandular trichomes. Additionally, the non-glandular trichomes play the prominent roles in growth and development of plants.  This study was aimed to detect different types of trichomes on the leaf epidermal surface of S. macrosiphon, and their variations among different Iranian populations. Material and methods: Eight natural populations of this plant were harvested from diverse habitats in Iran. Plant samples were identified, according to the morphological descriptions are available in the  valuable references. Three flowering plants were selected from each population and one mature and intact leaf was obtained per individual. The leaves were fixed in fixative (F.A.A) solution for 48 h. The hand-made cuttings of leaf's blade were double-stained with methylene blue and carmine colors. Then, the thin slices of each population were examined using a light microscopy (Olympus CH2, Japan) at different magnifications. Results: the leaf epidermal surfaces were covered by a dense indumentum, which were composed of the glandular and non-glandular trichomes. The non-glandular trichomes had the simple and unbranched structures with one to five linear-arranged cells. The non-glandular trichomes had two cell types: long and short. However, the more frequent non-glandular trichomes were the long two to four-celled types. Meanwhile, in  Amir kabir population,  the five-celled non-glandular trichomes also had a high density. The glandular trichomes were detected as capitate, digitate, and peltate types. Two types of capiate trichomes were observed on the leaves surfaces: short-stalked and long-stalked capitate. The main difference between these types relates to the cell number of trichomes stalk. However, the short-stalked capitate trichomes were the dominant glandular type in all the populations, except for Arak and Mashhad populations, which had the peltate trichome as a more frequent form.  Conclusion: the capitate and peltate trichomes do not have the same ability to maintain the biosynthesized essential oil in their cellular structures. The stored essential oil in the capitate trichomes seeps out through the micropores in their apical cells. Since, the short-stalked capitate was the dominant form in most populations, the secretion of essential oil makes this species very fragrant. But, these plants are less fragrant in populations that have a higher number of peltate trichomes. The non-glandular hairs play a key role in protecting the plant from herbivore insects and the ultraviolet rays of the sun light. Moreover, they protect the leaf epidermal surface from extreme heat and cold. The type and density of other glandular and non-glandular trichomes widely differed among the populations which explored their adaptive importance in this species.

Abstract Introduction: Cartilage, a tissue without blood vessels and nerves, possesses inherently limited regenerative capacity following injury, often leading to progressive joint degeneration and conditions like osteoarthritis (OA) if left untreated. Current clinical interventions, such as surgical microfracture or autologous chondrocyte implantation (ACI), face significant challenges, including donor site morbidity, immune rejection, and the formation of fibrocartilage with inferior biomechanical properties. These limitations underscore the urgent need for novel therapeutic strategies that can effectively stimulate hyaline cartilage regeneration. In this context, mesenchymal stem cell-derived exosomes (MSC-Exos) have garnered attention as a cell-free regenerative tool, leveraging their cargo of bioactive molecules (e.g., miRNAs, cytokines, and growth factors) to modulate chondrogenesis, suppress inflammation, and enhance extracellular matrix (ECM) synthesis. Concurrently, glucosamine, a natural amino sugar and precursor for glycosaminoglycan (GAG) biosynthesis, has demonstrated dual functionality in joint health: not only does it serve as a building block for proteoglycans critical to cartilage integrity, but it also exhibits chondroprotective effects by mitigating ECM degradation and promoting stem cell chondrogenic differentiation. The potential synergy between MSC-Exos and glucosamine could thus address multiple facets of cartilage repair, combining anabolic stimulation (via exosomal signaling) with metabolic support (via glucosamine supplementation), offering a promising combinatorial approach to halt OA progression and restore functional cartilage.
Aims: This study aimed to investigate the combined effect of mouse bone marrow stem cell-derived exosomes and glucosamine on the expression of cartilage-specific genes, including Sox9, Acan, Col2a1, and Col10a1.
Materials and Methods: Bone marrow mesenchymal stem cells were prepared from NMRI mice. The mice were euthanized by cervical dislocation, the femoral heads were removed, and the bone marrow contents were transferred into a cell culture flask using a syringe containing culture medium. The bone marrow cells were cultured and were ready for use after 3 to 5 passages. The cell supernatant was separated, and exosomes were extracted from it by successive rounds of centrifugation followed by ultracentrifugation. Mesenchymal stem cell viability and determining the appropriate concentration of exosomes and glucosamine were performed using the MTT assay. The experiments were performed on mesenchymal stem cells in 4 groups: control, exosome, glucosamine, and exosome + glucosamine. The effects of exosomes and glucosamine on the expression of Sox9, Acan, Col2a1, and Col10a1 genes in mesenchymal stem cells were investigated in the presence of chondrogenic medium.
Results: According to the MTT assay results demonstrating the synergistic effect of exosomes and glucosamine, the combined concentrations of 15 μg/mL exosomes and 25 μg/mL glucosamine were chosen for subsequent applications. Real-time PCR results showed that the expression of Sox9, Acan, and Col2a1 genes in stem cells treated with exosomes and glucosamine significantly increased compared to the other groups after 14 days, while the expression of the Col10a1 gene significantly decreased compared to the other groups.
Discussion: The combined treatment of bone marrow–derived mesenchymal stem cell (BMSC) exosomes and glucosamine significantly upregulated the expression of key chondrogenic markers, including Sox9, Acan, and Col2a1, while downregulating the hypertrophic marker Col10a1. This gene expression profile suggests a dual beneficial effect: (1) promotion of chondrogenic differentiation and extracellular matrix (ECM) synthesis, and (2) suppression of hypertrophic differentiation, a critical factor in preventing cartilage calcification and osteoarthritis progression. These findings highlight the synergistic potential of BMSC exosomes and glucosamine as a combinatorial therapy for cartilage regeneration. By enhancing anabolic processes (Sox9-mediated chondrogenesis and aggrecan/collagen II deposition) and concurrently inhibiting catabolic pathways (Col10a1-associated hypertrophy), this strategy may offer a promising approach to delay or reverse early-stage cartilage degeneration in degenerative joint diseases
Conclusion: Our study reveals that combining bone marrow stem cell-derived exosomes with glucosamine synergistically enhances chondrogenesis by upregulating key cartilage markers (Sox9, Acan, Col2a1) while suppressing hypertrophy-related Col10a1. This dual action suggests that exosomes promote cartilage matrix synthesis through their bioactive cargo (e.g., miRNAs/growth factors), while glucosamine likely inhibits hypertrophic differentiation, potentially via modulation of the Wnt/β-catenin pathway. These findings support this combination as a promising strategy for improving cartilage repair and preventing OA progression, though further in vivo validation is needed.

Abstract Aim: Cartilage defects of the knee such as osteoarthritis disease (OA) are one of the most debilitating and public diseases that are related to high individual and socioeconomic problems. Many recent studies have applied mesenchymal stem cells (MSCs) incorporated with tissue engineering to repair articular cartilage defects or regeneration of OA.  So, the choice of the best cell type in this regard is one of the challenging issues of tissue engineering and OA cell therapy. This study aims to find the best cell source for the regeneration of critical size defects of cartilage knee using autologous chondrocytes and bone marrow-derived MSCs (Bm-MSCs), and adipose-derived MSCs (Ad-MSCs) that were isolated, cultured, and expanded in similar in vitro conditions.
Materials and Methods: In the current study, Najdi sheep were used at 12 months of age. After standard anesthesia, cartilage was isolated from the hyaline cartilage at the end of the ribs. The Bm-MSCs and Ad-MSCs were isolated from bone marrow and tails' adipose tissue, respectively. Following the enzymatic digestion of cartilage and adipose tissues, using collagenase I enzymes, the chondrocyte, Ad-MSCs, and Bm-MSCs were cultured in growth media at 37º C with similar conditions. Then, MSCs were identified by morphology analysis and also osteogenic/adipose/chondrogenic differentiation, in vitro. In addition, chondrocytes were identified by morphology and analysis of cartilage-related gene expression such as Aggrecan, Col II, and SOX9 genes by Real-time PCR technique. After that, an amount of 5×10⁶ cells/ml from each cell source was seeded in the type I collagen gel and transplanted into an experimentally created articular cartilage defect in the knee’s sheep model. Two months after transplantation, the animals were sacrificed in the standard ways and the implanted tissue was removed. The range of regeneration was investigated by macroscopic scoring and histological staining such as H&E and safranin o/fast green.
Results: MSCs showed spindle shape of morphology, and skeletal differentiation were confirmed the identity of MSCs and chondrocytes. The macroscopic observation showed that the defects in cell-treated groups of chondrocytes, Bm-MSCs, and Ad-MSCs were filled with hyaline cartilage-like tissue in contrast to the control groups of untreated and sham (without cell) groups. In addition, the surface of new cartilage formed in Bm-MSCs and chondrocyte groups appeared to be smoother than in the Ad-MSCs group and the hyaline cartilage of Bm-MSCs is more clearly than that of the Ad-MSCs group. Although histological scores (ranging from 1 to 4) were evaluated, there was no significant difference among the three experimental groups regarding newly formed cartilage repair tissues. Furthermore, the histological analysis of H&E and safranin O revealed that all defects were filled by chondrocyte-like cells that were enclosed in the secreted matrix (*P<0.05).
Conclusion: We used three prevalent and main autologous cell sources such as chondrocyte, BM-MSCs, and AD-MSCs cells in exactly equal conditions to find the most significant cell sources for critical size defect of cartilage in sheep’s knees. The results demonstrated that three cell sources are suitable for this purpose; Although the Ad-MSCs due to ease and more accessibility are further recommended.

Abstract Introduction: Di(2-ethylhexyl) phthalate (DEHP) is a commonly used plasticizer in polyvinyl chloride (PVC) products, including medical devices such as syringes, IV tubing, blood bags, respiratory circuits, and dialysis equipment. Due to its weak physical bonding to the PVC matrix and lack of covalent interactions, DEHP can leach into biological fluids upon contact. This becomes particularly concerning during prolonged clinical exposure, as it allows the compound to enter the bloodstream and potentially affect sensitive tissues, including bone marrow. Bone marrow-derived mesenchymal stem cells (BMSCs) are multipotent progenitors with the ability to differentiate into osteoblasts, playing a vital role in bone remodeling and regeneration. Given their sensitivity to environmental toxins, DEHP exposure represents a significant risk to BMSC viability and osteogenic function. Evidence suggests that DEHP disrupts osteogenesis by inducing oxidative stress and downregulating essential genes involved in matrix formation and mineralization. Curcumin (Cur), a bioactive polyphenol extracted from the rhizome of Curcuma longa, possesses strong antioxidant, anti-inflammatory, and cytoprotective properties. It is nontoxic, affordable, and widely available, making it a promising candidate for counteracting xenobiotic-induced oxidative damage. This study explores whether Curcumin can mitigate DEHP’s deleterious effects on BMSCs during osteogenic differentiation.
Aim: We hypothesize that co-treatment with Cur will ameliorate DEHP-mediated disruption in cell viability, antioxidant capacity, and osteogenic marker expression.
Materials and Methods: BMSCs were isolated from adult male Wistar rats under sterile conditions and cultured to passage three. Cells were divided into four experimental groups and exposed for 21 days to the following treatments: (1) control, (2) DEHP (100 μM), (3) Curcumin (0.1 μM), and (4) DEHP + Curcumin. The cells were cultured in osteogenic differentiation medium throughout the treatment period. Cell viability was assessed using the tetrazolium-based MTT assay. Osteogenic differentiation was evaluated via Alizarin Red staining for mineral deposition, calcium quantification, and alkaline phosphatase (ALP) activity measurement. Oxidative stress was assessed by quantifying intracellular malondialdehyde (MDA) levels, a lipid peroxidation marker. Antioxidant enzyme activity for catalase (CAT) and superoxide dismutase (SOD) was measured spectrophotometrically. Total antioxidant capacity (TAC) was evaluated using commercial kits. For molecular analysis, total RNA was extracted from cells and reverse-transcribed into complementary DNA (cDNA). Semi-quantitative PCR was performed to measure expression levels of osteogenic differentiation-related genes: Smad1, Bmp2, Bmp7, Runx2, Alp, Col-1A1, and Osteocalcin (Oc). Gapdh served as the internal control. Data were statistically analyzed using ANOVA with Tukey’s post-hoc test. A threshold of p < 0.05 was considered statistically significant.
Results DEHP treatment caused a marked reduction in BMSC viability (p < 0.0001), confirming its cytotoxicity. Cells co-treated with Curcumin showed a significant restoration in viability (p < 0.01), indicating Cur’s protective effects. Alizarin Red staining revealed diminished extracellular matrix mineralization in DEHP-treated cells, with corresponding reductions in calcium content and ALP activity. Notably, Curcumin co-treatment restored all markers to levels comparable to those seen in control cells. ALP, an early osteogenic marker, showed complete recovery, underlining Cur’s efficacy in preserving bone-forming potential. Gene expression analysis demonstrated that DEHP downregulated osteogenic genes (Smad1, Bmp2, Bmp7, Runx2, Col-1A1, Oc). Co-treatment with Curcumin significantly reversed this suppression, elevating transcript levels to near control values. These results suggest a transcriptional rescue linked to improved redox homeostasis. Oxidative stress measurements indicated that DEHP increased MDA levels while suppressing CAT and SOD activity, as well as total antioxidant capacity (p < 0.0001). Curcumin treatment effectively reduced MDA concentrations (p < 0.05), and boosted CAT and SOD activity (p < 0.01). TAC was significantly elevated in Curcumin-treated groups (p < 0.0001), indicating improved redox balance and defense against oxidative damage.
Conclusion: This study provides strong evidence that DEHP impairs BMSC viability and osteogenic differentiation primarily through oxidative stress mechanisms. Curcumin co-treatment mitigates DEHP-induced cellular damage, restores osteogenic function, and enhances antioxidant defenses. These findings highlight Curcumin’s potential as a therapeutic agent to counteract phthalate toxicity in clinical contexts involving prolonged exposure to DEHP-containing materials.

Abstract Introduction: Okra (Abelmoschus esculentus (L.) Moench) is an annual herbaceous plant belonging to the family Malvaceae, valued for its high nutritional and medicinal properties. It is cultivated widely in various tropical and subtropical regions of the world, including many areas of Iran, where it serves as an important vegetable crop in local diets and traditional medicine. Aim: This study investigated the fatty acid composition and antioxidant activity of seed oils extracted from seven cultivars of Okra, namely White, Sultani, Red, Velvet, Green, Texas, and Fawn.
Material and methods: Mature and intact seeds from each okra cultivar were powdered, and their oils were extracted using a modified Folch et al. method. The fatty acid methyl esters (FAMEs) were prepared from the extracted oils through transesterification. The resulting FAMEs were analyzed using gas chromatography (GC) and gas chromatography–mass spectrometry (GC–MS) apparatus to determine their fatty acid profiles. The antioxidant activity of the extracted oils was evaluated using the DPPH radical scavenging assay. Data were statistically analyzed using SPSS and MVSP software.
Results: The seed oils of all okra cultivars contained both saturated and unsaturated fatty acids, with total proportions varying among the cultivars. The highest total saturated fatty acid content (42.53%) was observed in the White cultivar, while the lowest (36.41%) occurred in the Sultani cultivar. Palmitic acid was the predominant saturated fatty acid in all cultivars, with the highest (35.74%) and lowest (30.70%) levels detected in the White and Texas cultivars, respectively. Stearic acid was the second most abundant saturated fatty acid, ranging from 3.69% (Sultani) to 4.94% (White). Trace saturated fatty acids, including heptadecenoic, arachidic, behenic, and lignoceric acids were detected in all cultivars, whereas lauric acid was only found in a few cultivars (Velvet, Texas, and White cultivars). Unsaturated fatty acids constituted 57.33–63.33% of the total oil composition, with the lowest proportion in the White cultivar and the highest in the Sultani cultivar. Linoleic acid was the major unsaturated fatty acid in all cultivars except White, where oleic acid predominated. The highest linoleic acid content (11.40%) was recorded in the Red cultivar, and the lowest (8.25%) in White cultivar. In contrast, oleic acid content ranged from 18.20% (Red) to 49.30% (White). Trace amounts of other unsaturated fatty acids, including myristoleic, palmitoleic, trans-heptadecenoic, linolenic, gondoic, and erucic acids, were also detected across all cultivars. Additionally, eicosadienoic acid was present in trace amounts in all cultivars except White cultivar. Phytochemical analyses using the UPGMA dendrogram and Principal Component Analysis (PCA) grouped the cultivars into two main clusters, indicating significant phytochemical diversity. The predominance of oleic acid in the White cultivar, in contrast to linoleic acid in the others, along with the high palmitic acid content in the Velvet and White cultivars, contributed to the distinct placement of the White, Green, and Velvet cultivars from the remaining ones. The antioxidant activity, evaluated using the DPPH radical scavenging assay, revealed IC₅₀ values ranging from 689.23 µg/mL (Green cultivar) to 2551.12 µg/mL (Texas cultivar).
Discussion: The variation in fatty acid composition among the seven Okra cultivars reflects significant biochemical diversity. Palmitic acid was the main saturated fatty acid in all cultivars, while oleic and linoleic acids dominated the unsaturated fractions. The White cultivar, with its high oleic acid level, exhibited greater oil stability potential, whereas other cultivars rich in linoleic acid offer higher nutritional value. Differences in fatty acid profiles may be related to cultivar-specific desaturase activity. The antioxidant activity varied notably, with the Green cultivar showing the strongest and the Texas cultivar the weakest radical scavenging capacity, likely due to variations in phenolic and unsaturated compounds. The UPGMA and PCA analyses confirmed chemical diversity and separated White, Green, and Velvet cultivars based on their distinct fatty acid compositions.

Abstract Introduction: About half of the causes of infertility are related to men. The sperm of infertile men often have different functional and structural defects. Among these defects is sperm DNA damage, which can be caused by DNA fragmentation, improper chromatin packaging, and epigenetic defects. All men have some degree of damage to sperm DNA, but when the percentage of this damage increases significantly, it causes pregnancy disruption, miscarriage, and failure in assisted reproductive methods. Studies have shown that 15% of couples are unable to have children despite trying to conceive and are considered infertile, with half of these cases of infertility being due to male factors. The majority of male infertility is due to abnormal sperm. Therefore, these individuals are candidates for assisted reproductive techniques such as IVF and ICSI and may have tried these treatments repeatedly and failed. One of the most important factors in the success of in vitro fertilization is the health of sperm DNA. Various studies show that the lower the quality of sperm, the more problems sperm DNA health faces.
Aims: Therefore, the purpose of conducting research to identify sperm DNA damage and study its effect on the success rate of assisted reproductive methods, including microinjection, in infertile couples is essential, with the aim of improving sperm quality before starting the treatment cycle and imposing excessive costs on couples.

Materials and methods: The present study was experimental and appropriate laboratory equipment, materials, and solutions were used in the research process. In this experiment, semen samples were collected from 60 infertile couples (30 of which were considered positive control samples) who had referred to the Royesh Infertility Treatment Center in Karaj for ICSI treatment. Oligoasthenospermia samples and positive control samples were selected and separated. Anti-Mullerian hormone (AMH) of women candidates for ICSI, which is present in the patient's serum and measured through a blood test, was examined, and those with AMH equal to 2 and above were selected. Then, sperm washing was performed for intracytoplasmic injection. Sperm injection into the egg is usually done 2-3 hours after ovulation. In the next step, sperm DNA breakage was assessed. Failure assessment was performed immediately after sample receipt. The collected data was analyzed using SPSS software, which provides a summary of the methods and techniques used.
Results: The average embryo quality in the normal sperm group was 93.35%, while this index was only as high as 75.74% in the DNA-damaged sperm group. Also, the average sperm velocity in the normal sperm group was 36.2%, while this index in the DNA-damaged sperm group showed a small value of 5.53%. Also, The average percentage of non-motile sperm, sperm with DNA breakage was 82.17%, and in the normal sperm group, the average was 41.93%. In addition, in the DNA breakage sperm group, the average percentage of sperm with rotational movement was 17%, sperm with slow progressive movement was 5.4%, and sperm with fast progressive movement was 0.1%, while in the normal sperm group, the desired indicators showed averages of 0.87%, 25.13%, and 11.07%, respectively. The results of the study also showed that sperm with DNA breakage has a significant impact on reducing embryo quality. And sperm DNA damage can severely and significantly reduce sperm velocity and count. And finally, sperm DNA breakage has a significant impact on sperm shape.
Discussion: The results obtained indicate that DNA breakage is more common in oligoasthenospermic individuals than in healthy individuals, and this breakage significantly affects embryo quality.
Conclusion: Since in assisted reproductive methods, damage to sperm DNA may cause treatment failure, it is recommended to check the sperm DNA fragmentation rate (SDFA) before choosing the appropriate treatment method. And in many cases, if there is damage to sperm DNA, appropriate treatment can improve sperm quality. SDFA testing is a reliable method for assessing sperm DNA health, which greatly contributes to the clinical diagnosis and treatment of male infertility and is of great value in the success of treatment methods.

Abstract Introduction: Prostate cancer is one of the most prevalent malignancies among men worldwide and represents a major cause of cancer related morbidity and mortality. Despite advances in screening and therapeutic approaches, resistance to standard treatments, including androgen‑deprivation therapy, remains a major clinical challenge in prostate cancer. As a result, identifying novel compounds that can suppress proliferation or modulate key molecular regulators of the cell cycle is of considerable interest. Papaverine is an isoquinoline alkaloid derived from the opium poppy, widely known for its long‑standing clinical use as a smooth‑muscle relaxant and vasodilator. In recent years, it has gained attention for its potential anticancer effects, including its influence on mitochondrial function, cellular energy metabolism, and the modulation of signaling pathways involved in cell cycle regulation. However, the specific effects of papaverine on prostate cancer cells have not yet been clearly elucidated. Tamoxifen, a selective estrogen receptor modulator widely used in breast cancer therapy, has also demonstrated off target antiproliferative effects in various tumor models.
Aims: The present study aimed to evaluate the effects of papaverine and tamoxifen on the viability and molecular regulatory profile of DU145 prostate cancer cells line. Specifically, the study investigated the cytotoxic potential of these compounds on different concentrations and incubation times, and their ability to alter the expression of CDK4 gene, a key mediator of G1 to S phase progression, along with two microRNAs, miR-146a and miR-22, known to influence cell cycle control and tumorigenic pathways. By integrating cellular and molecular findings, the study sought to clarify whether these compounds could serve as potential modulators of prostate cancer cell growth.
Materials and Methods: DU145 human prostate cancer cell line were cultured under standard conditions and treated with different concentrations of papaverine and tamoxifen. Cell viability was assessed for 24, 48, and 72 hours using the MTT assay (3 [4,5 dimethylimidazole-2-yl] 2,5 diphenyl tetrazolium bromide). Dose–response curves were generated to calculate IC50 values for each compound at the specified time points. To investigate molecular changes, total RNA was extracted from the treated and the control cells, and the expression levels of CDK4 gene, miR-146a, and miR-22 were quantified using Real Time PCR with appropriate internal controls. Relative expression changes were determined using the 2-ΔΔCt method. The data were analyzed using SPSS software. Results were presented as Mean ± SD, and significance level of p ≤ 0.05 was considered statistically significant.
Results: Treatment of DU145 prostate cancer cell line with various concentrations of papaverine and tamoxifen resulted in a significant reduction in cell viability in a concentration and time dependent manner, as measured by the MTT assay for 24, 48, and 72 hours. IC50 values were successfully determined for both compounds across the different time points.
Molecular analysis using Real Time PCR showed that treatment with either papaverine or tamoxifen led to a marked downregulation of CDK4 expression compared with untreated controls. In contrast, both compounds induced a significant up‑regulation of miR‑146a and miR‑22 levels.
Discussion: The observed decrease in viability suggests that papaverine and tamoxifen exert notable antiproliferative effects on DU145 prostate cancer cells. The downregulation of CDK4 gene provides a mechanistic link to cell cycle arrest, as CDK4 is a central regulator of G1‑to‑S phase progression. Such suppression is in line with reduced cellular proliferation and supports the therapeutic potential of these compounds.
The increased expression of miR‑146a and miR‑22 further highlights the involvement of tumor suppressive microRNAs in mediating these effects. Both miRNAs have been implicated in regulating pathways associated with proliferation, apoptosis, and oncogenic signaling. Their upregulation may contribute to the inhibition of cell growth through negative modulation of critical oncogenic targets.
These results align with prior evidence demonstrating tamoxifen’s antiproliferative effects but also importantly emphasize that the molecular and cellular impacts of papaverine in prostate cancer remain poorly understood. The current findings provide initial insight into papaverine’s interaction with cell cycle regulatory genes and microRNAs, suggesting a potential role in prostate cancer growth inhibition. Considering the adverse effect profile associated with tamoxifen, papaverine may therefore represent a more favorable candidate for further investigation, particularly as a potential alternative with a potentially safer therapeutic window.

Abstract Introduction
Cancer remains one of the leading causes of mortality worldwide, and gastric cancer is among the most aggressive malignancies because of its high incidence, late diagnosis, and poor prognosis. Conventional treatments such as surgery, radiotherapy, and chemotherapy are often associated with systemic toxicity, severe side effects, and drug resistance. Cisplatin (CDP), a widely used chemotherapeutic drug for gastric cancer, induces apoptosis through DNA damage; however, its clinical use is limited by toxicity and resistance. Nanotechnology has enabled the development of smart nanocarriers that improve drug stability, bioavailability, and targeted delivery. Cellulose nanocrystals (CNCs) are attractive drug carriers because of their biodegradability, mechanical strength, and controlled-release properties. In addition, bovine serum albumin (BSA), polyvinyl alcohol (PVA), polyethylene glycol (PEG), and hyaluronic acid (HA) improve biocompatibility and targeting efficiency through interactions with CD44 receptors overexpressed on gastric cancer cells.
Aims
This study aimed to design and evaluate a smart nanocarrier system composed of CNCs, BSA, PVA, PEG, and HA for targeted cisplatin delivery to AGS gastric cancer cells. The study investigated physicochemical properties, pH-responsive drug release, cytotoxicity, selectivity toward cancer cells, and apoptosis induction to determine the therapeutic potential of the synthesized nanocapsules.
Materials and Methods
Cellulose nanocrystals were extracted from raw cellulose using sulfuric acid hydrolysis (64%), followed by ultrasonication, centrifugation, and dialysis. Cisplatin was loaded onto CNCs at different concentrations, and drug-loading efficiency was measured spectrophotometrically at 370 nm. Multifunctional BSA-PVA-CNC-CDP nanoparticles coated with PEG-hyaluronic acid were synthesized through sequential coating and hydrogel formation processes. First, CNC-CDP complexes were coated with bovine serum albumin and stabilized in a PVA hydrogel matrix. The nanoparticle surface was then functionalized with NH2-PEG-HA using EDC/NHS coupling chemistry to enhance targeting ability and biological stability. The synthesized nanoparticles were characterized using XRD, TGA, and DLS analyses to evaluate crystalline structure, thermal stability, particle size, and zeta potential. Drug release behavior was investigated at pH 7.4 and pH 5.8 to simulate physiological and tumor microenvironments. Cytotoxicity and biocompatibility were assessed in AGS gastric cancer cells and normal GES-1 cells using the MTT assay. The selectivity index (SI) was calculated to evaluate preferential toxicity toward cancer cells. In addition, apoptosis induction was analyzed using Annexin V/PI staining and flow cytometry. Statistical analysis was performed using one-way ANOVA followed by Duncan’s multiple range test.
Results
XRD analysis demonstrated that all synthesized nanoparticles and nanocapsules exhibited predominantly amorphous structures, confirming successful incorporation of CNC-CDP complexes into the BSA-PVA matrix. TGA results showed that increasing CNC concentration from 50 to 200 mg significantly improved the thermal stability of CNC-CDP nanoparticles. Furthermore, BPC(100 mg)CP-HA nanocapsules displayed greater structural stability and lower thermal degradation compared with drug-free nanocapsules. DLS analysis indicated that increasing CNC concentration increased particle size from 322 to 363 nm while reducing zeta potential from +13.05 to +2.14 mV. Surface coating with BSA, PVA, PEG, and HA further increased particle size to 476 nm, confirming the formation of multilayered nanocapsules. Drug release studies demonstrated significantly higher cisplatin release under acidic conditions (pH 5.8) compared with physiological conditions (pH 7.4). In contrast, increasing CNC concentration reduced the release rate because of the formation of denser polymeric networks.
Biological evaluations showed that drug-free nanocapsules exhibited negligible toxicity against both AGS and GES-1 cells, confirming favorable biocompatibility. In contrast, cisplatin-loaded nanocapsules significantly reduced AGS cell viability. Among all formulations, BPC(50 mg)CP-HA demonstrated the strongest anticancer activity, with an IC50 value of 224.75 µg.mL⁻¹ and the highest selectivity index (SI=1.75). Flow cytometry analysis further revealed that this formulation induced the highest levels of apoptosis in AGS cells, whereas free cisplatin predominantly caused necrotic cell death.
Conclusion
The BPCCP-HA smart nanocapsules developed in this study represent a promising strategy for targeted gastric cancer therapy. The nanocarrier system improved physicochemical stability, enabled pH-responsive controlled release of cisplatin, enhanced selective toxicity toward AGS cancer cells, and promoted apoptosis-mediated cell death while maintaining low toxicity toward healthy cells. These findings suggest that the designed nanocarrier system may effectively overcome major therapeutic limitations associated with conventional cisplatin treatment.