Cell and Tissue Journal

Abstract Bladder cancer is one of the most common malignancies of the urinary tract and ‎originates from the bladder epithelium. Despite advances in surgery, chemotherapy, ‎radiation, and immunotherapy, bladder cancer remains a major clinical challenge ‎because of its high recurrence rate, risk of progression, and the need for long-term ‎surveillance and repeated interventions. In recent years, vaccine-based immunotherapy ‎has gained increasing attention as a promising strategy for improving antitumor ‎immune responses, reducing relapse, and enhancing therapeutic outcomes across ‎different disease stages.‎
Bacillus Calmette–Guérin (BCG) is the first and most established vaccine used in ‎bladder cancer treatment and remains the standard intravesical adjuvant ‎immunotherapy for non muscle invasive bladder cancer (NMIBC). Its antitumor ‎activity is largely mediated by immunomodulatory effects within the bladder ‎microenvironment, including activation of innate immune cells, induction of ‎pro inflammatory cytokines, and subsequent priming of adaptive immune responses ‎that support cytotoxic T cell–mediated tumor control. However, a substantial ‎proportion of patients experience intolerance, inadequate response, recurrence, or ‎resistance, underscoring the need for next generation vaccine strategies with improved ‎specificity and durability.‎
Accordingly, several emerging vaccine platforms have been investigated. Cell based ‎vaccines, particularly dendritic cell (DC) vaccines, aim to exploit the antigen presenting ‎function of DCs to enhance tumor specific T cell activation using tumor lysates, ‎defined tumor associated antigens, or engineered immunostimulatory constructs. In ‎parallel, monoclonal antibody–based approaches that target tumor associated antigens ‎have been explored as immunotherapeutic tools that may complement vaccine-‎induced immunity. Peptide vaccines provide a more defined strategy by stimulating ‎antigen specific responses against selected epitopes, and their performance may be ‎strengthened through the use of appropriate adjuvants, optimized epitope selection, ‎and improved delivery systems. More recently, mRNA vaccines have attracted ‎considerable interest because they enable rapid and flexible design, can encode one or ‎multiple tumor antigens, and may induce robust cellular immunity. These platforms ‎also offer opportunities for personalization based on patient specific antigenic profiles ‎and may be manufactured in a scalable manner.‎
Combination therapy is an important direction in this field. Pairing vaccine approaches ‎with immune checkpoint inhibitors (e.g., PD 1/PD L1 blockade) may enhance ‎therapeutic efficacy by reversing tumor mediated immune suppression and improving ‎the magnitude and durability of antitumor responses. In addition, integrating vaccines ‎with conventional modalities such as chemotherapy or radiotherapy may further ‎augment immunogenicity through increased antigen release, activation of danger ‎signaling pathways, and immune priming, thereby potentially converting ‎immunologically “cold” tumors into more responsive disease states.‎
This review summarizes major vaccine strategies investigated for bladder cancer, ‎including BCG, cell based vaccines, peptide vaccines, and mRNA vaccines, and ‎discusses their proposed mechanisms, advantages, and current limitations. Despite ‎encouraging progress, key challenges remain, including tumor heterogeneity, antigen ‎loss, immune evasion, optimization of delivery and dosing schedules, manufacturing ‎complexity, and cost. Future research should focus on refining antigen discovery and ‎vaccine design, identifying predictive biomarkers for patient selection and response ‎monitoring, optimizing combination regimens to maximize synergy, and conducting ‎larger, well designed clinical studies to support clinical translation. Overall, vaccine-‎based immunotherapy represents a rapidly evolving and promising avenue that may ‎contribute to more effective, durable, and individualized treatment of bladder cancer ‎for patients across disease stages.‎

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: Sperm cryopreservation is a critical biotechnology for genetic resource conservation and artificial insemination in domestic animals. However, conventional glycerol-based methods cause severe damage to the plasma membrane, mitochondria, and sperm DNA due to intracellular ice crystal formation, osmotic stress, and increased reactive oxygen species (ROS), ultimately reducing post-thaw viability and fertility. The studies included in this meta-analysis cover multiple economically important species, including cattle, sheep, goats, and particularly poultry (roosters), all of which hold significant value for both economic productivity and biodiversity conservation in Iran. This systematic review and meta-analysis synthesizes evidence from 11 comparative studies to evaluate the efficacy of optimized cryopreservation protocols (e.g., DMA pellets, dry thawing, nano-enhanced extenders, and programmable freezing) versus standard glycerol-based methods on sperm viability and motility.
Aims: The primary objective is to quantify the pooled effect size of optimized cryopreservation methods specifically on post-thaw viability and progressive motility compared to standard glycerol‑based protocols. The secondary objectives are to identify sources of heterogeneity, to assess publication bias, to provide predictions for future studies, and to offer practical, actionable recommendations for implementing these technologies in Iran, particularly for poultry gene banking and the preservation of native breeds.
Materials and Methods: Following PRISMA 2020 guidelines, we systematically searched PubMed, Scopus, Web of Science, Google Scholar, SID, MagIran, and reference lists up to November 2025. Search keywords included: sperm cryopreservation, motility, DMA, nanoparticles, and poultry.
Inclusion criteria were: comparative studies in domestic animals, reporting mean ± SD for post-thaw viability/motility, with a sample size of at least 6 (n ≥ 6). Quality assessment of the included studies was performed using a risk-of-bias tool. Data were analyzed using CMA v4 with a random-effects model. Effect size was calculated as the standardized mean difference (SMD). Heterogeneity was assessed using the statistic and Q-test. Publication bias was evaluated via Funnel Plot and Egger's test.
Results: Eleven studies (comprising 276 sperm samples) were included in the final meta-analysis. Optimized methods significantly improved viability (SMD = 4.514, 95% CI: 3.253–5.776, Z = 7.014, p < 0.001; I² = 86.2%) and motility (SMD = 3.161, 95% CI: 2.378–3.943, Z = 7.919, p < 0.001; I² = 75.7%). The Q-test also revealed significant heterogeneity (p < 0.001), which is entirely expected given the diversity of animal species and cryoprotectant types employed across the primary studies. However, Egger's test indicated no significant publication bias for either outcome (p > 0.05).
Discussion: The pooled analysis showed very large effect sizes favoring optimized methods for both viability and motility. The high heterogeneity was expected and partly explained by species differences and the type of cryoprotectant used. From a biological perspective, DMA offers distinct advantages over glycerol due to its lower molecular weight and faster membrane permeability, which reduces equilibration time and minimizes cytotoxicity. Programmable freezing, by precisely controlling the cooling curve, prevents thermal shock and preserves the structural integrity of the spermatozoa.
Moreover, the incorporation of nanoparticles (e.g., selenium or titanium dioxide) into extenders effectively scavenges ROS, thereby inhibiting lipid peroxidation and optimizing sperm energy metabolism. The dry-thawing method further reduces cold-shock injury by minimizing the time spermatozoa spend in the critical temperature zone (−15 to 0°C), thus preventing recrystallization damage. The findings strongly support the adoption of DMA pellets, dry thawing, and nano‑enhanced extenders in breeding programs and gene banks, especially in Iran.
Conclusion: Optimized protocols substantially outperform standard methods. Given Iran's diverse climatic conditions and the urgent need to protect native poultry breeds from environmental and epidemiological threats, implementing these advanced protocols in national genetic resource centers and the country's gene bank is essential. Furthermore, standardizing rapid dry-thawing protocols across artificial insemination centers nationwide would be a highly effective step toward increasing practical fertility rates, reducing sperm wastage in commercial flocks, and ultimately supporting national food security goals. Therefore, we recommend the widespread adoption of these optimized methods to enhance fertility outcomes and ensure robust genetic preservation in domestic animals.

Abstract Objective:
Exosomes are nano-sized extracellular vesicles secreted by various cell types, playing a pivotal role in intercellular communication and the regulation of numerous biological processes, including inflammation, angiogenesis, and tissue regeneration. In recent years, mesenchymal stem cell (MSC)-derived exosomes have attracted considerable attention due to their ability to transfer bioactive molecules such as proteins, lipids, and RNAs to target cells, thereby modulating cellular functions without the risks associated with direct stem cell transplantation. On the other hand, low-frequency electromagnetic fields (LF-EMF) have been independently reported to enhance cell proliferation, migration, and wound closure through mechanisms involving calcium signaling and growth factor expression. Despite the individual therapeutic potentials of exosomes and LF-EMF, the synergistic effects of their combination on skin cells, particularly keratinocytes which are central to re-epithelialization during wound healing, have remained largely unexplored. Therefore, this experimental study was designed to investigate the combined effect of bone marrow stem cell-derived exosomes and LF-EMF on key cellular responses of keratinocytes, including apoptosis and the expression of inflammatory and anti-inflammatory genes.
Materials and Methods:
Bone marrow mesenchymal stem cells (BM-MSCs) were isolated from healthy adult rats and cultured under standard conditions until reaching 70–80% confluence. The cell culture supernatant was collected, and exosomes were isolated using a well-established ultracentrifugation protocol involving sequential centrifugation steps at 300×g, 2000×g, and 10,000×g to remove cell debris and large vesicles, followed by ultracentrifugation at 100,000×g for 70 minutes at 4°C. The resulting pellet, enriched with exosomes, was characterized in terms of size, morphology, and specific surface markers using dynamic light scattering (DLS), transmission electron microscopy (TEM), and Western blotting for CD63 and CD81. For the main experiment, keratinocytes (HaCaT cell line) were cultured and divided into four groups: control (untreated), exosome-treated alone, LF-EMF treated alone (exposed to a 50 Hz, 1 mT electromagnetic field for 1 hour daily for three consecutive days), and a combination group receiving both exosomes and LF-EMF under identical conditions. Following treatment, the rate of apoptosis among keratinocytes was quantitatively assessed using the Annexin V-FITC/PI double-staining assay analyzed by flow cytometry. Additionally, total RNA was extracted from each group, reverse transcribed into cDNA, and subjected to real-time PCR to measure the expression levels of five target genes: transforming growth factor beta 1 (TGFβ1), interleukin-18 binding protein (IL-18bp), interleukin-1 beta (IL-1β), interleukin-18 (IL-18), and tumor necrosis factor-alpha (TNF-α). Relative gene expression was calculated using the ΔΔCt method with GAPDH as the housekeeping gene.
Results:
The flow cytometric analysis of Annexin V staining revealed that the lowest percentage of apoptotic keratinocytes was observed in the combination group (exosomes + LF-EMF), indicating a significant protective synergy compared to either treatment alone. The reduction in apoptosis was statistically significant (p < 0.05) relative to control and individual treatment groups. Moreover, real-time PCR data demonstrated a remarkable upregulation of the anti-inflammatory and tissue-repair-related genes in the combination group. Specifically, the expression of IL-18bp, a natural inhibitor of IL-18-mediated inflammation, and TGFβ1, a key cytokine involved in extracellular matrix deposition and wound contraction, increased significantly compared to the other groups (p < 0.01). Conversely, the combination treatment led to a marked downregulation of pro-inflammatory cytokines: TNF-α, IL-1β, and IL-18 gene expression levels were substantially reduced compared to the control and single-treatment groups. These molecular changes indicate a shift from a pro-inflammatory to an anti-inflammatory, pro-healing environment within the keratinocyte population.
Conclusion:
The findings of this study provide strong evidence that the combined application of BM-MSC-derived exosomes and low-frequency electromagnetic fields exerts protective and immunomodulatory effects on keratinocytes. The significant reduction in apoptosis, coupled with the favorable modulation of inflammatory gene expression (upregulation of TGFβ1 and IL-18bp; downregulation of TNF-α, IL-1β, and IL-18), suggests that this dual strategy enhances cell survival and reduces excessive inflammation, both of which are critical for efficient wound healing. These results highlight the potential of combining exosome therapy with physical stimuli like LF-EMF as a novel, cell-free regenerative approach. Future studies should focus on identifying the specific molecular cargo within exosomes responsible for these synergistic effects and evaluating the efficacy of this combination in in vivo wound models. Overall, the exosome-LF-EMF combination represents a promising candidate for developing advanced wound healing therapies, especially for chronic or non-healing wounds.

Abstract Introduction: Catharanthus roseus (Madagascar periwinkle) is a vital medicinal plant renowned for producing anticancer terpenoid indole alkaloids (TIAs), including vincristine and vinblastine. However, the inherently low yield of these high‑value metabolites (typically <0.001% dry weight) necessitates effective elicitation strategies. Methyl jasmonate (MJ) and salicylic acid (SA) are well‑known elicitors that activate plant defense responses and secondary metabolism through distinct but interconnected signaling pathways (COI1‑dependent jasmonate signaling and NPR1‑mediated SA pathway). While individual effects of MJ and SA have been reported, their synergistic potential on a comprehensive set of physiological, biochemical, and molecular parameters in C. roseus has not been systematically investigated.
Aims: This study aimed to (1) evaluate the individual and combined effects of MJ and SA on biomass, membrane stability index (MSI), anthocyanin, chlorophyll (SPAD), protein content, antioxidant enzyme activities (SOD, CAT, GPX), essential oil (EO) yield, and the expression of four key TIA biosynthetic genes (TDC, STR, SGD, and DAT); and (2) identify the optimal concentration combination for maximizing alkaloid production.
Material and Methods: A factorial experiment based on a completely randomized design (CRD) with three replications was conducted. Four‑month‑old C. roseus 'Pink Rose' plants were foliar‑sprayed with MJ (0, 0.5, 1, 2 mM) and SA (0, 1, 2, 3 mM), individually or in combination, at 15‑day intervals. The treatments were applied three times to ensure sustained elicitation, and all plants were maintained under controlled greenhouse conditions (16 h light/8 h dark, 25±2°C day/20±2°C night, 60±5% relative humidity). Sampling was performed 24 h after the final treatment. Physiological traits (fresh/dry weight, MSI, anthocyanin, SPAD, total protein) and antioxidant enzyme activities were measured using standard spectrophotometric methods. Total protein content was determined by the Bradford assay, and antioxidant enzyme activities (SOD, CAT, and GPX) were assayed following established protocols. Essential oil was extracted by hydrodistillation from dried leaf powder and quantified gravimetrically. Gene expression was analyzed by qRT‑PCR using 18S rRNA as the internal reference. Data were subjected to two‑way ANOVA and Duncan's multiple range test (p ≤ 0.05) using SAS 9.2.
Results: The interaction between MJ and SA was significant (P ≤ 0.01) for all measured traits. Two combined treatments (1 mM MJ + 3 mM SA and 2 mM MJ + 2 mM SA) were identified as the optimal consensus treatments, as they ranked in the highest statistical groups across the majority of measured traits. Fresh weight and dry weight reached their highest values under 1 mM MJ + 2 mM SA, with increases of approximately 47% and 64% over the control, respectively. MSI improved by ~50% under both optimal treatments. Protein content increased by ~2.9-fold, while activities of SOD, CAT, and GPX rose approximately 3-fold compared to the control. Anthocyanin accumulation peaked (~2.5-fold over control) under 2 mM MJ + 3 mM SA, with 1 mM MJ + 3 mM SA placing in the same statistical group. Essential oil yield increased approximately 2-fold under the optimal combined treatments. All four TIA pathway genes (TDC, STR, SGD, and DAT) were upregulated by 2.4- to 3.8-fold, with STR showing the greatest induction (~3.8-fold) and SGD the least (~2.2–2.4-fold). Higher concentrations of SA (3 mM) in combination with lower MJ levels, or certain high-dose combinations, produced diminished responses for some traits, indicating concentration-dependent optima.
Discussion: These findings indicate that simultaneous MJ and SA elicitation activates coordinated physiological and transcriptional responses, with 1 mM MJ + 3 mM SA and 2 mM MJ + 2 mM SA as consensus optima. Enhanced antioxidant capacity, membrane stability, essential oil yield, and TDC/STR/SGD/DAT expression support a synergistic, concentration-dependent effect that may improve anticancer alkaloid production. Future field-scale validation and direct vinblastine/vincristine quantification are needed.
Conclusion: Co‑application of MJ and SA induces a strong combined effect, enhancing antioxidant capacity, essential oil accumulation, and transcriptional activation of TIA biosynthetic genes. This effect offers a possible approach to boost the production of anticancer alkaloids in C. roseus. Future field‑scale validation and direct quantification of vinblastine/vincristine are recommended.

Abstract Background and Objective: Antimicrobial resistance (AMR) is an important environmental and public health concern, and aquatic sediments can serve as reservoirs of antibiotic resistance genes (ARGs). Increasing evidence indicates that non-antibiotic contaminants, including heavy metals and pesticides, may also be associated with the environmental resistome. Zinc (Zn) is an essential trace element, and azinphos-methyl is an organophosphorus insecticide that may enter aquatic ecosystems through agricultural runoff. The southern Caspian Sea receives inputs from agricultural, industrial, and urban sources, and its coastal sediments provide a suitable environment for the accumulation of chemical contaminants and diverse microbial communities. However, limited information is available concerning the simultaneous spatial association of Zn and azinphos-methyl with ARGs and bacterial community structure in this region.

Aims: This study investigated the associations of Zn and azinphos-methyl with ARG abundance and bacterial community structure in coastal sediments of the southern Caspian Sea using shotgun metagenomic analysis.

Materials and Methods: Eight coastal sampling stations were selected (n = 8). Genomic DNA was extracted from 300 mg of sediment, and shotgun metagenomic libraries were sequenced on the Illumina NovaSeq 6000 platform. ARGs were identified using CARD and ResFinder, and ARG abundance was normalized as reads per million (RPM). Bacterial taxonomic composition was characterized at the phylum level. Zn concentrations were determined by ICP-MS, and azinphos-methyl was quantified using GC-MS. Alpha diversity was assessed using the Shannon index, and beta diversity was assessed using principal coordinates analysis (PCoA). Associations were evaluated using Spearman rank correlation analysis, with P < 0.05 considered statistically significant.

Results: Proteobacteria was the dominant bacterial phylum across most stations, reaching up to 94.4% relative abundance. The Shannon diversity analysis indicated that L1 and L2 had the highest diversity, whereas L8 showed the lowest diversity, with PCoA demonstrating separation of L8 from other stations. Zn was measurable at only two stations (342.5 mg/kg at L1 and 122.76 mg/kg at L3). Azinphos-methyl was detected at three stations, with the highest concentration at L4 (0.463 ng/g). Spearman correlation analysis identified eight ARGs significantly and positively associated with Zn, with qacA showing the strongest association (ρ = 0.990, P < 0.001). Azinphos-methyl showed significant positive correlations with 25 ARGs, with bacA showing the strongest association (ρ = 0.892, P = 0.0029). At the bacterial phylum level, Zn showed four significant positive associations (SAR324 clade, Margulisbacteria, LCP-89, and Spirochaetota), whereas azinphos-methyl showed a significant negative association with Bacteroidota (ρ = −0.810, P = 0.015).

Discussion: The observed correlations indicate that spatial variation in Zn and azinphos-methyl concentrations was accompanied by differences in the abundance of several ARGs. The strongest Zn–ARG association was observed for qacA, a gene associated with multidrug efflux systems. Azinphos-methyl showed significant positive associations with a larger number of ARGs than Zn, with the strongest association observed for bacA. The bacterial community analysis provided an additional ecological context for the observed ARG patterns. However, such mechanisms cannot be directly inferred from the observational metagenomic data generated in the present study. The interpretation of the findings should particularly consider the limited sample size and the distribution of the investigated contaminants.

Conclusion: This study investigated the spatial associations of Zn and azinphos-methyl with ARGs and bacterial community structure in coastal sediments of the southern Caspian Sea. Zn showed significant positive correlations with eight ARGs, with qacA showing the strongest association (ρ = 0.990, P < 0.001). Azinphos-methyl showed significant positive correlations with 25 ARGs, with bacA showing the strongest association (ρ = 0.892, P = 0.0029). At the bacterial phylum level, Zn was positively associated with SAR324 clade, Margulisbacteria, LCP-89, and Spirochaetota, whereas azinphos-methyl showed a significant negative association with Bacteroidota. Overall, the findings demonstrate statistical co-variation among non-antibiotic contaminants, ARG abundance, and bacterial community structure. However, the observational design, the small number of independent samples (n = 8), and the highly uneven distribution of Zn and azinphos-methyl require cautious interpretation. The identified relationships should be considered exploratory and hypothesis-generating candidate associations rather than evidence of causal effects.

Abstract Introduction: Liver diseases cause ~2 million deaths annually; cholestasis, defined as impaired bile formation or flow, can progress to cirrhosis and liver failure. Excess Cu2+ catalyzes Fenton chemistry, generating hydroxyl radicals that induce lipid peroxidation (elevated malondialdehyde, MDA) and reduce glutathione (GSH). The resulting oxidative stress activates nuclear factor-kappa B, releasing pro-inflammatory cytokines (TNF-α, IL-6, IL-1β). Concurrently, copper disrupts BCL-2 family balance, increasing Bax, releasing cytochrome c, activating caspase-3, and executing mitochondrial apoptosis. Critically, Cu also suppresses canalicular transporters BSEP (bile salt export pump) and MDR3 (phosphatidylcholine floppase), causing intrahepatic accumulation of toxic bile acids and a vicious cholestatic cycle.
Current therapies (ursodeoxycholic acid, obeticholic acid) have limited efficacy and side effects. The polyphenol-rich plant extracts offer multi-target hepatoprotection. Black-peel pomegranate (Punica granatum L. var. Black), an Iranian variety, has exceptionally high anthocyanin and ellagitannin content.
Aims: This study evaluated the hepatoprotective effects of aqueous black-peel pomegranate peel extract (BPPE) against CuSO4-induced cholestatic liver injury in rats, examining serum cholestatic markers (ALP, GGT), hepatic oxidative stress (MDA, GSH), copper accumulation, and mRNA expression of inflammatory (NF-κB, TNF-α, IL-6, IL-1β), apoptotic (Bax, Bcl-2, cytochrome c, caspase-3), and canalicular transporter (BSEP, MDR3) genes.
Materials and methods: Thirty-six male Wistar rats (190-220 g) were divided into six groups (n=6): (1) Control (normal water); (2) CuSO4 (150 mg/kg/day in drinking water); (3) CuSO4 + vehicle (1% CMC gavage); (4-6) CuSO4 + BPPE at 50, 100, and 150 mg/kg/day by gavage. Treatments lasted 30 days. At the endpoint, blood and liver were collected. Serum ALP and GGT were measured using commercial kits. Hepatic MDA (Thiobarbituric acid method), GSH (DTNB assay), and copper content (atomic absorption spectrometry after acid digestion) were quantified. Relative mRNA expression of target genes was analyzed by real-time PCR with SYBR Green, normalized to GAPDH. Data (mean ± SEM) were compared by one-way ANOVA and Tukey test.
Results: CuSO4 significantly increased serum ALP (~3-fold) and GGT (~6-fold) vs. control. BPPE at all doses lowered these, with 100 mg/kg restoring levels to near-control. Hepatic MDA doubled, and GSH fell to 38% of control in the CuSO4 group. BPPE (100 mg/kg) normalized MDA and recovered GSH to ~80% of control, not differing from healthy animals. Copper content in liver increased 3.3-fold with CuSO4; 100 mg/kg BPPE reduced it by ~54%, reaching a level similar to controls.
Gene expression (100 mg/kg BPPE vs. CuSO4 alone): NF-kB was upregulated 3.32-fold by CuSO4 and normalized to 1.04-fold by BPPE. IL-6 (3.7-fold) reduced to 1.36-fold; IL-1β (2.4-fold) to 1.8-fold; TNF-α (1.75-fold) to 1.5-fold. The pro-apoptotic Bax, cytochrome c, and caspase-3 were upregulated 2.7-, 1.6-, and 1.2-fold by CuSO4, while anti-apoptotic Bcl-2 dropped to 0.33-fold. BPPE decreased Bax, cytochrome c, and caspase-3, and increased Bcl-2 to 1.5-fold above control, restoring the Bax/Bcl-2 ratio. BSEP and MDR3 were suppressed by CuSO4 to 0.51- and 0.43-fold of control; BPPE partially restored them to 0.79- and 0.82-fold, respectively.
Discussion: The extract normalization of ALP and GGT confirms anti-cholestatic efficacy. CuSO4-induced oxidative stress (MDA rise, GSH depletion) was counteracted by BPPE’s direct radical scavenging and metal chelation, evidenced by a 54% reduction in hepatic copper. The observed bell-shaped dose-response (slight decline at 150 mg/kg) may reflect the pro-oxidant behavior of excess polyphenols.
At the molecular level, BPPE nearly abolished NF-κB activation and downstream cytokines, breaking the inflammatory feed-forward loop. This anti-inflammatory effect likely contributed to the partial restoration of BSEP and MDR3 expression, as these transporters are suppressed by cytokines and positively regulated by FXR. By preserving canalicular transporters, the extract aids bile acid secretion and interrupts the cholestatic cycle. Concomitantly, BPPE suppressed the mitochondrial apoptotic cascade, lowering Bax, cytochrome c, and caspase-3 while upregulating Bcl-2, an effect attributable to its abundant anthocyanins. Therefore, BPPE acts through an integrated polypharmacology: chelating copper, scavenging free radicals, blocking NF-κB, inhibiting apoptosis, and reviving bile transporter expression.
Conclusion: Aqueous black-peel pomegranate peel extract at 100 mg/kg effectively protects against CuSO4-induced cholestatic liver injury by reducing oxidative stress, chelating copper, suppressing NF-kB-driven inflammation, blocking mitochondrial apoptosis, and partially restoring BSEP/MDR3 transporters. These findings identify this extract as a safe, cost-effective natural candidate for managing heavy-metal-associated cholestatic liver damage. Further studies on chronic models and clinical validation are warranted.

Abstract During the past 30 years, researchers have made a remarkable progress in identifying the biological (bacteria, viruses), biochemical (chemical compounds) and biophysical (ionizing radiation) cause of human cancer. The term ˝Cancer˝ refers to 277 forms of cancer diseases.
Scientists have determined the process of cancer formation from a consequence of accumulating multiple mutations in human genome. These genetic disruptions would eventually change the normal process of cellular proliferations and differentiation.
The genetic alterations are frequently indicative of poor prognosis for most human cancers.
Both nonhereditary and hereditary cancers are caused by genetic disorders that change the cellular growth control system.
 Genes associated with human cancer formation include four classes of genes: 1. Oncogenes (which increasing their activities end to uncontrollable growth of cells), 2. Tumor suppressor genes, 3. DNA repairing genes, 4. Apoptotic genes.
Over activated oncogenes which cause cellular proliferation. In contrast, inactivated tumor suppressor genes lose their inhibitory effect which is crucial to prevent inappropriate growth. DNA repairing proteins fix the damage and apoptotic proteins cause the pre-cancer cell to commit suicide.
We have over millions of genes in each somatic cell of our body. After sequencing all human genome in 2003, we noticed that Only 23,500 genes are active which encode over 400,000 proteins needed for physiological functions.
99.9% of genome is identical in all humans worldwide. Only 0.1% of the whole genome differ which cause the genetic variations.
Up to 93% of all human cancers are non-hereditary and the remaining 7% are hereditary. A wealth of information has been indicated by the potential use of bioinformatics and molecular techniques for cancer screening, prognosis and monitoring of the efficacy of anticancer therapies. In recent years, molecular genetics have greatly increased our understanding of the basic mechanisms in cancer development. The essential outcome of these molecular studies is that the cancer can be considered as the genetic disease.
 

Abstract Aim: Pentoxifylline (PTX) is a methylxanthine derivative medicine used to improve motility of human spermatozoa in-vitro. It is commonly used in treatment of male-factor infertility, including asthenozoospermia. This study aimed to evaluate the effect of PTX on human sperm parameters and DNA integrity from asthenozoospermic problem.
Material and methods: A total of 38 infertile men with asthenozoospermia were allocated in this experimental study.  Specimens were randomly divided into experimental group treated with 3.6 mM PTX, and control group. All samples were incubated at 37˚ C for 45 min. Semen parameters and sperm DNA fragmentation were measured using sperm chromatin dispersion (SCD) test.
Results: PTX improved sperm motility, significantly, compared to the control (85.76±5.96 Vs 79.44±9.37, respectively, p < .01). There was also a significant decrease in sperm viability in the PTX- treated group in comparison to controls (87.7±8.3 Vs 83.5±9, respectively, p < .01). In addition, sperm DNA fragmentation was higher in PTX-treated group compared to control (23.36±10.25 and 18.5±8.74, respectively, p < .0001).
Conclusion: PTX might have some negative impact(s) on sperm DNA quality, although it has improved the sperm motility. Further studies are needed to elucidate the safety of PTX treatments in ART clinics.

Abstract Plants produce a big group of secondary metabolites which are used as medicinal compounds. According to recent estimates, global market value of herbal medicines, including medicinal plants and their products, significantly has been increasing. Considering to the fact that most of the world market for medicinal plants, production and supply of secondary metabolites derived from these plants are concerned and the plant secondary metabolites are of high economic value. Chemical synthesis of these metabolites is an expensive process. So production of metabolites by different biotechnological methods such as cell culture is a useful alternative. Molecular recognition and elicitor-plant receptors interaction is a complex process requiring for signal transduction. Biotic elicitors induce secondary metabolites and hypersensitive responses by activation of defense mechanisms. Manipulation of cell culture media by elicitors is an important strategy for inducing secondary metabolism and production of valuable metabolites. Molecular recognition and elicitor-plant receptors interaction is a complex process requiring for signal transduction. Following perception of elicitor signals, rapid defense responses can be organized as follows: increase of ionic currents across the plasma membrane, reactive oxygen species (ROS) production, activation of defense gene expression, structural changes in the cell wall and phytoalexin production. In this study, different aspects of increasing the production of secondary metabolites in cell culture of plants by biotic elicitors is investigated.  

Abstract Aim: The aim of this study was to investigate the effect of vitamin E on sperm parameters in adult rat treated with para-nonylphenol.
Material and methods: Adult male rats were divided into four groups: Control, para-nonylphenol, vitamin E and para-nonylphenl+vitamin E. Oral treatments were performed till 56 days. At the end of treatments, body and left testis weight were recorded and left caudal epididymis was cut in a medium. Released spermatozoa were used to analyze sperm parameters such as sperm number, viability, morphology and motility. Sperm chromatin quality was assessed by nuclear staining using acridine orange and aniline blue.
Results: Body and testis weight as well as normal sperm morphology showed no significant change in four groups. A significant decrease in the number, viability and motility of the sperm was found in rats treated by para-nonylphenol compared to the control. This decrease was significantly compensated by vitamin E in para-nonylphenol+vitamin E group compared to para-nonylphenol group. The application of vitamin E alone could significantly increase sperm viability and motility as comared with the control. Para-nonylphenol had no effect on sperm DNA integrity and histon-protamine replacement compared to the control.
Conclusion: Vitamin E, as a potent antioxidant, could protect the adverse effect of para-nonylphenol on certain sperm parameters in adult rats.
 

Abstract Cryopreservation is the most effective method for long-term maintenance of sperm. There are several procedures for freeze-thawing of semen and each may impose damage on sperm function, viability and finally decreases semen quality and fertility potential. In addition to decreased percentage of sperm viability and motility after freeze-thawing, percentage of DNA damage is also increases due to high level of oxidative stress. To minimize these damages, we need to increase our insights regarding different cryopreservation procedures, cryoprotectant and antioxidant supplements, which can protect sperm membrane during cryopreservation. Therefore, by using these experiments, we can improve the efficiency of these procedures. In this review, we discuss about principles of cryopreservation, types of freeze-thawing methods, advantages and disadvantages of each of these methods, effects of freezing on sperm parameters and clinical outcomes, and finally role of antioxidants in preservation of sperm integrity during freeze-thawing. For this review, all relevant information was collected via databases such as PubMed and Google Scholar during the period of 1966-2017.
 

Abstract Aim: This study was done to evaluate the therapeutic effects of Curcuma longa L. rhizome powder and bovine ghee mixture on experimental stomach ulcers in rat.
Material and methods: Stomach ulcer induction was done in 48 food deprived male rats (250-300 gr) using orally Indomethacin (50 mg/kg suspended in 1% carboxymethyl cellulose. Animals divided randomly into five groups. The normal and untreated (Indomethacin) group just received normal saline. The treated groups received different doses of C. longa and bovine ghee (500 and 750 mg C. longa in 10 ml ghee/per kg) for three successive days. On the third day, the rats were killed and their stomachs were removed for histological studies. Stomach ulcers number and length were measured and also ulcer index was calculated.
Results: Results showed that the mixture of C. longa and bovine ghee is significantly caused decreasing gastric ulcer index, inflammatory cells, blood capillaries densities (p < 0.001) and increasing mucosal layer thickness (p < 0.001) and secreted mucus (p < 0.05) in treated groups in comparison with untreated group.
Conclusion: According to the results, the mixture of C. longa and bovine ghee is significantly accelerated healing of experimental stomach ulcers.

Abstract Aim: The main goal was to evaluate the impact of different incubation time intervals on human sperm DNA status using terminal deoxyribonucleotidyl transferase–mediated dUTP nick-end labeling (TUNEL) test.
Material and methods: This prospective study involved 21 normozoospermic specimens. After direct swim-up, sperm cells were incubated at 37^°C and DNA damage was evaluated at different time intervals (0, 1, 2 and 3 h).  After slide fixation with methanol 100% for 4 minutes and rinse in phosphate-buffered solution (PBS), TUNEL was added to each slide and incubated for 60 minute in dark. Eosin-Nigrosin and Papanicolaou staining protocols were applied in order to assess sperm viability and morphology, respectively.
Results: Sperm viability and normal morphology was improved after sperm processing (100%, 72.3% respectively, p < 0.0001). The rate of DNA damage was significantly higher after 2h compared to 0h (9.19±0.8% Vs 4.9±0.9%, respectively, p=0.008). Also there was significant difference in abnormal sperm DNA between 3h and 1h (10.95±0.7% Vs 7.1±1%, respectively, p=0.020).
Conclusion: Incubation of prepared normozoospermic samples at 37^°C more than 2 h may be associated with sperm DNA fragmentation. Therefore, it seems that incubation of human spermatozoa at 37^°C should be limited up to 2h prior to use in ART clinics. 

Abstract Angiogenesis is a necessary process in many physiological and pathological events such as tumor growth. Therefore angiogenesis therapy can be a strategy to cancer treatment. It is important to identify different aspects of tumor’s angiogenesis.  Formation of functional vessels within tumors is essential for its growth and progression. The induction of new blood vessel growth by tumors is mediated through different molecules. A balance between pro-angiogenic and anti-angiogenic growth factors strictly control the process. This fact has led to design of therapeutic agents against tumor angiogenesis. Recently, endogenous (such as interleukin -8) and exogenous angiogenesis inhibitors (such as: Avastin) have been described. Furthermore because of the low mutagenic potential of endothelial cells, tumor’s vessel do not show resistance to the effects of many of these compounds. In addition, it is recognized that anti-angiogenesis and anti-cancer tumors can be more effective by combination of factors than the use of either alone. This article would review different aspects of angiogenesis in tumors. Since tumor growth is dependent on the development of blood vessels in the tumor, inhibition of angiogenesis maybe considered an appropriate treatment for cancer.