Cell and Tissue Journal

Abstract Introduction: Concerns about fossil energy costs, environmental deterioration, and energy security has created strong motivation for the research and development of routes to provide sustainable and renewable fuels. In recent years, the use of biomass to produce highly valued chemicals has attracted widespread attention. Lignocellulosic biomass, as a promising renewable resource for biofuel production, has distinct advantages in terms of economic and environmental benefits. The conversion of renewable raw materials to hydrocarbon fuels is an attractive alternative to fossil fuels from economic and environmental perspectives. The production process of lignocellulosic biomass mainly consists of biomass accumulation, biomass decomposition, simple sugars, and conversion of sugars to biofuel. One of the crucial steps for the economic success of lignocellulosic biofuels depends on the inhibition of competitive metabolism in microorganisms to achieve high productivity. To date, there has been a growing focus on the use of S. cerevisiae and E. coli as cell lines. These two cellular factories have well known advantages. They are genetically transmissible and several tools are available for genetic manipulation. In order to produce xylonate, the engineered xylose is first converted by a dehydrogenase into the intermediate xylonolactone, which is then slowly converted to xylonate in a nonenzymatic reaction.
Aim: The organic compound D-1,2,4-Butanetriol (BT) is a valuable chemical with wide-ranging applications in various fields such as pharmaceuticals, paper, polymer materials, and military applications. However, the chemical synthesis routes for BT have many drawbacks. By genetically modifying microorganisms, the metabolic pathway for producing many substances, including BT, can be engineered. When D-xylose is supplied to the bacterium, it is first converted into an intermediate compound called xylonolactone. This compound slowly converts into xylonate through a non-enzymatic reaction. To produce xylonate, the engineered bacteria receive xylose, which is initially converted by a dehydrogenase reaction catalysed by the xylose dehydrogenase enzyme into an intermediate compound, xylonolactone. Xylonolactone is slowly converted to xylonate in a nonenzymatic reaction. Xylonate is a five-carbon organic acid. Over the past few years, xylonate has increasingly been considered as an important chemical due to its potential as an important chemical component. Xylonate has many applications in the food, chemical, and pharmaceutical industries. Specifically, xylonate can act as a precursor for the synthesis of D-1,2,4-Butanetriol and as a concrete water reducing agent. E. coli was chosen as the target strain for genetic and metabolic engineering due to its fast growth in inexpensive culture media, the presence of two enzymes for BT synthesis, and product formation in less than 24 hours of fermentation. This study aimed to clone and express xylose dehydrogenase from Caulobacter vibrioides in E.coli.
Materials and Methods: At first, to access the bacterial gene sequence, the genome of the target bacterium was extracted. Then, to create a strain expressing the enzymes xylose dehydrogenase and xylonolactonase, the genes for these proteins were amplified from Caulobacter vibrioides CB1 and transferred into E. coli. For this purpose, the target genes were amplified using specifically designed primers via the Polymerase Chain Reaction (PCR) method and initially cloned into a pTZ57cloning vector and then subcloned into pET 26b expression vector. At the final step, the expression of the enzyme was assessed by SDS-PAGE, and the other confirmation was the reduction of NAD+ to NADH, which was used as an activity indicator of the enzyme, as investigated by a change in NADH absorbance at 340 nm.
Results: Confirmatory tests were performed to ensure the presence of the gene in the vectors (using restriction enzymes and colony PCR for gene amplification). The expression and activity of the enzyme were analyzed. The recombinant protein's presence was confirmed by SDS-PAGE for the xylose dehydrogenase gene, with a molecular weight of 52.2 kDa. The estimated expression level of the recombinant protein was approximately 25%.
Conclusion: The objective of this research was solely to establish the metabolic pathway for xylonate production in E. coli by surface expression of enzymes in this pathway (xylose dehydrogenase). The results obtained in this study confirm that half of the pathway is active at the cell surface, but further experiments are required to determine the precise production levels and complete the pathway.

Abstract Introduction: Global climate change has significantly increased the frequency and intensity of abiotic stresses, thereby limiting plant growth, development, and overall yield by damaging physiological systems. In response to environmental stressors, plants have evolved various adaptive mechanisms, including the accumulation of compatible solutes such as glycine betaine (GB), which plays a pivotal role in protecting cellular functions under adverse conditions. Also known simply as betaine, this compound is a methylated glycine derivative recognized across plant species for its ability to mitigate the deleterious effects of stressful environments. Its zwitterionic structure; comprising a positively charged trimethylammonium group and a negatively charged carboxyl group, confers high solubility and chemical stability to the molecule.
Owing to its excellent biocompatibility, favorable carbon-to-nitrogen ratio, and high-concentration accumulation, glycine betaine can enhance plant tolerance against a wide spectrum of abiotic stresses. Specifically, GB contributes to photosynthetic recovery and the alleviation of oxidative stress by reducing the accumulation and facilitating the detoxification of reactive oxygen species (ROS). Furthermore, it plays a crucial role in stabilizing membranes and macromolecules, while protecting key components of the photosynthetic apparatus, such as the Rubisco enzyme, Photosystem II (PSII), quaternary enzymes, and complex protein structures. Notably, glycine betaine can accumulate at high concentrations within plant cells without interfering with normal metabolic processes, thereby significantly increasing resilience to various osmotic stresses, extreme temperatures (heat and cold), and oxidative damage. The biosynthesis of glycine betaine occurs through distinct metabolic pathways, including the choline oxidation pathway (prevalent in plants and mammals), the direct glycine methylation pathway (specific to certain bacteria and halophytes), the choline dehydrogenase pathway, and the serine metabolism pathway. Such diversity underscores the vital importance of this osmolyte in mediating responses to environmental stresses. Recently, biotechnological interventions, such as Agrobacterium-mediated transformation, have successfully enhanced stress tolerance in susceptible species by overexpressing key genes, most notably codA, BADH, GSMT, and SDMT. Given the functional diversity of genes involved in the glycine betaine biosynthetic pathway, extensive efforts have been made to develop transgenic plants capable of effective accumulation of this metabolite; however, serious challenges such as unstable and weak transgene expression remain as key obstacles in this path. Factors including promoter type, genomic integration site, and epigenetic factors can influence the final performance. Furthermore, the overexpression of enzymes in the glycine betaine biosynthetic pathway may potentially impair growth by disrupting metabolic stability. Therefore, future research should focus on decoding the molecular networks regulating the biosynthesis, signaling, and transport of glycine betaine, particularly its crosstalk with phytohormones, transcription factors, and the identification of stress-inducible promoters. Additionally, optimizing transformation protocols and synchronizing glycine betaine gene expression with the overall plant metabolism are essential.
Aims: This review examines the biosynthesis, physiological functions, and molecular regulation of glycine betaine (GB). It highlights genetic engineering techniques to boost GB production, offering sustainable strategies for enhancing crop tolerance to environmental stresses.
Conclusion: In-planta biosynthesis of glycine betaine (GB) offers a more sustainable alternative to exogenous application, aligning closely with the principles of green agriculture. By integrating GB synthesis pathways, genetically engineered crops can autonomously boost metabolite production and bolster stress resilience, thereby eliminating the logistical costs of external treatments. Furthermore, a more profound investigation into these biosynthetic pathways will facilitate the identification and cloning of novel target genes, ultimately maximizing GB accumulation and enhancing environmental tolerance.

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.