Significant Changes In Expression Research Articles

Enhancing Pseudomonas syringae pv. Actinidiae sensitivity in kiwifruit by repressing the NBS-LRR genes through miRNA-215-3p and miRNA-29-3p identification

Kiwifruit bacterial canker, caused by Pseudomonas syringae pv. actinidiae (PSA), poses a grave threat to the global kiwifruit industry. In this study, we examined the role of microRNAs (miRNAs) in kiwifruit’s response to PSA. Kiwifruit seedlings subjected to PSA treatment showed significant changes in both miRNA and gene expression compared to the control group. We identified 364 differentially expressed miRNAs (DEMs) and 7170 differentially expressed genes (DEGs). Further analysis revealed 180 miRNAs negatively regulating 641 mRNAs. Notably, two miRNAs from the miRNA482 family, miRNA-215-3p and miRNA-29-3p, were found to increase kiwifruit’s sensitivity to PSA when overexpressed. These miRNAs were linked to the regulation of NBS-LRR target genes, shedding light on their role in kiwifruit’s defence against PSA. This study offers insights into the miRNA482-NBS-LRR network as a crucial component in enhancing kiwifruit bioresistance to PSA infestation and provides promising candidate genes for further research.

Jamaican fruit bat (Artibeus jamaicensis) insusceptibility to mucosal inoculation with SARS-CoV-2 Delta variant is not caused by receptor compatibility

The ancestral sarbecovirus giving rise to SARS-CoV-2 is posited to have originated in bats. While SARS-CoV-2 causes asymptomatic to severe respiratory disease in humans, little is known about the biology, virus tropism, and immunity of SARS-CoV-2-like sarbecoviruses in bats. SARS-CoV-2 has been shown to infect multiple mammalian species, including various rodent species, non-human primates, and Egyptian fruit bats. We show that SARS-CoV-2 can utilize Jamaican fruit bat (Artibeus jamaicensis) ACE2 spike for entry in vitro. Therefore, we investigate the Jamaican fruit bat as a possible in vivo model to study reservoir responses. We find that SARS-CoV-2 Delta does not efficiently replicate in Jamaican fruit bats in vivo. We observe infectious viruses in the lungs of only one animal on day 1 post-inoculation and find no evidence of shedding or seroconversion. This is possibly due to host factors restricting virus egress after aborted replication. Furthermore, we observe no significant immune gene expression changes in the respiratory tract but do observe changes in the intestinal metabolome after inoculation. This suggests that, despite its broad host range, SARS-CoV-2 is unable to infect all bat species, and Jamaican fruit bats are not an appropriate model to study SARS-CoV-2 reservoir infection.

Effect of Triclisia subcordata Oliv. (Menispermaceae) leaves on hormonal imbalance and genes expression in the ovaries of letrozole-induced polycystic rats via in vivo and computational approaches

The hormonal imbalance, insulin sensitivity, and ovarian function are among the biological processes linked to polycystic ovarian syndrome (PCOS). Hence, in PCOS ovaries, the impact of Triclisia subcordata Oliv. leaf (TSL) on hormone levels and the expression of the genes FTO, CYP11A, IRS, P53, and HSD17B2 was studied. Comparable to a normal control group, the rats treated with TSL methanol extract had the lowest levels of testosterone and the highest quantity of the follicle-stimulating hormone. In the ovaries of PCOS rats, TSL methanol extract strongly reduced HSD17B2 expression and significantly increased IRS expression. Furthermore, significant changes in FTO expression were detected, suggesting possible correlations with metabolic disorders, hormone dysregulation, and features of ovarian function. Additionally, P53 expression significantly decreased. GC-MS analysis revealed the presence of farnesol formate, 4-(4-ethylcyclohexyl) -1-pentyl, and cyclohexene as phytoconstituents. In T. subcordata leaves, cyclohexene, 4-(4-ethylcyclohexyl)-1-pentyl-, and farnesol formate showed a more unusual and specific binding to the p53 core, FTO protein, and cytochrome P450 CYP11A1 domains. According to the molecular dynamics study, one of the main mechanisms that have been proposed for the recovery of metabolic syndrome in affected individuals is the stabilization of proteins by the binding of these chemicals to the targeted structure. All these results suggest that Triclisia subcordata leaves may have complex impacts on PCOS-related molecular pathways, including restoring normalcy to insulin sensitivity, androgen metabolism, hormone balance, and ovarian function.

Deletion of an evolutionarily conserved TAD boundary compromises spermatogenesis in mice.

Spermatogenesis is a complex process that can be disrupted by genetic and epigenetic changes, potentially leading to male infertility. Recent research has rapidly increased the number of protein coding mutations causally linked to impaired spermatogenesis in humans and mice. However, the role of non-coding mutations remains largely unexplored. As a case study to evaluate the effects of non-coding mutations on spermatogenesis, we first identified an evolutionarily conserved topologically associated domain (TAD) boundary near two genes with important roles in mammalian testis function: Dmrtb1 and Lrp8 . We then used CRISPR-Cas9 to generate a mouse line where 26kb of the boundary was removed including a strong and evolutionarily conserved CTCF binding site. ChIP-seq and Hi-C experiments confirmed the removal of the CTCF site and a resulting increase in the DNA-DNA interactions across the domain boundary. Mutant mice displayed significant changes in testis gene expression, abnormal testis histology, a 35% drop in the estimated efficiency of spermatogenesis and a 28% decrease in daily sperm production compared to littermate controls. Despite these quantitative changes in testis function, mutant mice show no significant changes in fertility. This suggests that non-coding deletions affecting testis gene regulation may have smaller effects on fertility compared to coding mutations of the same genes. Our results demonstrate that disruption of a TAD boundary can have a negative impact on sperm production and highlight the importance of considering non-coding mutations in the analysis of patients with male infertility.

Nematode Galectin Inhibits Basophilic Leukaemia RBL-2H3 Cells Apoptosis in IgE-Mediated Activation.

Mast cells are essential immune cells involved in the host's defence against gastrointestinal nematodes. To evade the immune response, parasitic nematodes produce a variety of molecules. Galectin 1, produced by Teladorsagia circumcincta (Tci-gal-1), reduces mast cell degranulation and selectively regulates mediator production and release in an IgE-dependent manner. To uncover the activity of Tci-gal-1, we have examined the effect of the protein on gene expression, protein production, and apoptosis in activated basophilic leukaemia RBL-2H3 cells. Rat RBL-2H3 cells were activated with anti-DNP IgE and DNP-HSA, and then treated with Tci-gal-1. Microarray analysis was used to examine gene expression. The levels of several apoptosis-related molecules and cytokines were determined using antibody arrays and ELISA. Early and late apoptosis was evaluated cytometrically. Degranulation of cells was determined by a β-hexosaminidase release assay. Treatment of activated RBL-2H3 cells with Tci-gal-1 resulted in inhibited apoptosis and decreased degranulation, although we did not detect significant changes in gene expression. The production of pro-apoptotic molecules, receptor for advanced glycation end products (RAGE) and Fas ligand (FasL), and the cytokines IL-9, IL-10, IL-13, TNF-α, and IL-2 was strongly inhibited. Tci-gal-1 modulates apoptosis, degranulation, and production of cytokines by activated RBL-2H3 cells without detectable influence on gene transcription. This parasite protein is crucial for modulation of the protective immune response and the inhibition of chronic inflammation driven by mast cell activity.

Vitamin D Receptor Regulates Liver Regeneration After Partial Hepatectomy in Male Mice.

Vitamin D signals through the vitamin D receptor (VDR) to induce its end-organ effects. Hepatic stellate cells control development of liver fibrosis in response to stressors and vitamin D signaling decreases fibrogenesis. VDR expression in hepatocytes, however, is low in healthy liver, and the role of VDR in hepatocyte proliferation is unclear. Hepatocyte-VDR null mice (hVDR) were used to assess the role of VDR and vitamin D signaling in hepatic regeneration. hVDR mice have impaired liver regeneration and impaired hepatocyte proliferation associated with significant differential changes in bile salts. Notably, mice lacking hepatocyte VDR had significant increases in expression of conjugated bile acids after partial hepatectomy, consistent with failure to normalize hepatic function by the 14-day time point tested. Real-time PCR of hVDR and control livers showed significant changes in expression of cell cycle genes including cyclins D1 and E1 and cyclin-dependent kinase 2. Gene expression profiling of hepatocytes treated with vitamin D or control showed regulation of groups of genes involved in liver proliferation, hepatitis, liver hyperplasia/hyperproliferation and liver necrosis/cell death. Together these studies demonstrate an important functional role for VDR in hepatocytes during liver regeneration. Combined with the known profibrotic effects of impaired VDR signaling in stellate cells, the studies provide a mechanism whereby vitamin D deficiency would both reduce hepatocyte proliferation and permit fibrosis, leading to significant liver compromise.

The effect of preoperative hormonal stimulation on the urethral plate; A histologic and histochemical study

Preoperative hormone stimulation (PHS) is used to increase the glans size and may improve the cosmetic appearance after hypospadias surgery. The exact effect of PHS on different penile tissues remains unclear and controversial. Previous studies showed that PHS increased vessel density in the foreskin. However, the effect of PHS on the urethral has never been studied before. In this study we examine the PHS effects on the urethral plate. Specimens of the urethral plate and the underlying tissue were excised to correct severe chordee in 16 children with proximal and perineal hypospadias with severe chordee in 2 groups. Group A consisted of 8 children received PHS prior to surgery. Group B included 8 children with proximal and perineal hypospadias with severe chordee who did not receive PHS and served as a control group. Specimens were examined blindly using hematoxylin-eosin and elastic van-Gieson stain as well as factor 8 and SMA antibodies. The median age of Group A was 13 (range 10-20) months. The median age of Group B (without prior PHS) was 17 (range 14-29) months (p=0,03). The median vessel-density in group A (82 vessels per mm2) was significantly higher than in Group B (65 per mm2), (p<0.05). The median vessel diameter was also significantly higher in Group A (13μm) than in Group B (11μm), (p<0.05). The median epithelial layer thickness in Group A was 110μm and in Group B 98μm, but showed no statistical significance (p=0,16). There was no significant change in dartos fascia layer thickness or androgen receptor expression. There was no visual change in the organization of elastic fibers. This study is the first study to document the effect of PHS on the urethral plate in hypospadias with severe chordee. PHS significantly increased the median vessel-density as well as median vessel diameter as compared to a matching control group. The findings of the study may justify PHS administration before the first operation to improve the vascularity and may reduce the severity of chordee.

Analysis of culture and RNA isolation methods for precision-cut liver slices from cirrhotic rats

Precision-cut liver slices (PCLS) are increasingly used as a model to investigate anti-fibrotic therapies. However, many studies use PCLS from healthy animals treated with pro-fibrotic stimuli in culture, which reflects only the early stages of fibrosis. The effects of different culture conditions on PCLS from cirrhotic animals has not been well characterized and there is no consensus on optimal methods. In this study, we report a method for the collection and culture of cirrhotic PCLS and compare the effect of common culture conditions on viability, function, and gene expression. Additionally, we compared three methods of RNA isolation and identified a protocol with high yield and purity. We observed significantly increased albumin production when cultured with insulin-transferrin-selenium and dexamethasone, and when incubated on a rocking platform. Culturing with insulin-transferrin-selenium and dexamethasone maintained gene expression closer to the levels in fresh slices. However, despite stable viability and function up to 4 days, we found significant changes in expression of key genes by day 2. Interestingly, we also observed that cirrhotic PCLS maintain viability in culture longer than slices from healthy animals. Due to the influence of matrix stiffness on fibrosis and hepatocellular function, it is important to evaluate prospective anti-fibrotic therapies in a platform that preserves tissue biomechanics. PCLS from cirrhotic animals represent a promising tool for the development of treatments for chronic liver disease.

P-400 Pregnancy-specific glycoproteins - potential markers for risk associated with preeclampsia during gestational SARS-CoV-2 Infection

Abstract Study question Could pregnancy-specific glycoproteins serve as potential markers for the risk of developing preeclampsia during gestational SARS-CoV-2 Infection? Summary answer Pregnancy-specific glycoproteins (PSGs) are significantly down-regulated in women with gestational SARS-CoV-2 infection, which could be used to diagnose high-risk pregnancies in assisted reproductive treatment (ART). What is known already In vitro fertilization (IVF) pregnancies are precious and face increased risks of preeclampsia. Similarly, SARS-CoV-2 infection during pregnancy increases the risk for preeclampsia. Precious pregnancies with ART along with SARS-CoV-2 infection can further exacerbate the risk for preeclampsia, affecting both maternal and fetal health. Thus, identifying molecular markers associated with preeclampsia risk in the context of SARS-CoV-2 infection is crucial for maternal and fetal health management, specifically in pregnancies with IVF. Study design, size, duration Placental samples (n = 16) were collected at term delivery from SARS-CoV-2 positive women (n = 7) and negative healthy women (n = 9). We studied separately, the gene expression pattern in maternal and fetal placental compartments. Interesting genes from RNA sequencing results were validated using real-time PCR. Participants/materials, setting, methods All pregnant women at their term delivery were clinically tested for SARS-CoV-2 infection by PCR. Maternal and fetal sides of placenta were collected in total RNA was extracted from the snap-frozen tissues. cDNA libraries were constructed using Smart-seq2 method and sequenced using Illumina NovaSeq 6000 sequencing platform. Bioinformatic analysis was using Partek® Flow® software, v9.0, DEGs were identified by applying FDR ≤ 0.05. qPCR was for validating the interesting genes. Main results and the role of chance Our analysis revealed 635 DEGs in the maternal placental tissue, with 518 upregulated and 117 downregulated genes in SARS-CoV-2-positive women compared with the healthy SARS-CoV-2-negative women. In contrast, the fetal compartment did not exhibit any significant changes in gene expression with SARS-CoV-2 infection. In the maternal compartment, we observed significant downregulation of nine genes belonging to the pregnancy-specific glycoprotein related to the immunoglobulin superfamily with active SARS-CoV-2 infection (fold change range from −13.70 to − 5.28; FDR ≤ 0.01). Additionally, comparing symptomatic women with healthy women, we identified 1788 DEGs. Furthermore, pathway enrichment analysis revealed that pathways related to oxidative phosphorylation, insulin secretion, cortisol synthesis, estrogen signaling, oxytocin signaling, and antigen processing were altered significantly in symptomatic women. Limitations, reasons for caution We have a limited sample size in this study, though we collected placental samples from all the women who were positive for SARS-CoV-2, at admission for delivery. Studies with larger sample sizes are warranted to further reconfirm the results. In vitro studies may help us to deepen our understanding. Wider implications of the findings This study sheds light on PSGs as potential biomarkers for risk for preeclampsia during gestational SARS-CoV-2 and reveals molecular pathways associated with an increased clinical risk of preeclampsia with COVID-19 during pregnancy. The findings offer valuable insights that could be further explored clinically in managing pregnant women with COVID-19. Trial registration number not applicable

Physiological and transcriptomic analysis reveals the toxicological mechanisms of polystyrene micro- and nano-plastics in Chlamydomonas reinhardtii

With the accumulation of plastic waste in the environment, the toxicity of micro- and nano-plastics (MNPs) to microalgae has attracted increasing attention. However, the underlying toxic mechanisms of MNPs remain to be elucidated. In this study, we synthesized micro- and nano-scale of polystyrene MNPs (PS MNPs) to investigate their toxicity and toxic mechanisms in Chlamydomonas reinhardtii. We found that PS MNPs significantly inhibit the production of photosynthetic pigments and increase soluble protein content. The detailed analysis of results shows that both materials affect photosynthetic efficiency by damaging the donor side, reaction center, and electron transfer of photosystem II. Moreover, compared to PS MPs, PS NPs have a greater negative impact on algal cells. Analyzing the transcriptome of cells suggests that the most sensitive metabolic pathways in response to PS MNPs involve oxidative phosphorylation, biosynthesis of secondary metabolites, and photosynthesis. Especially, genes related to photosynthesis and oxidative phosphorylation showed significant changes in expression after exposure to PS MNPs. This study provided molecular-level insights into the toxic mechanisms of PS MNPs on microalgae.

Decoding immune-related gene-signatures in colorectal neoplasia.

Colorectal cancer (CRC) is a significant health issue, with notable incidence rates in Norway. The immune response plays a dual role in CRC, offering both protective effects and promoting tumor growth. This research aims to provide a detailed screening of immune-related genes and identify specific genes in CRC and adenomatous polyps within the Norwegian population, potentially serving as detection biomarkers. The study involved 69 patients (228 biopsies) undergoing colonoscopy, divided into CRC, adenomatous polyps, and control groups. We examined the expression of 579 immune genes through nCounter analysis emphasizing differential expression in tumor versus adjacent non-tumorous tissue and performed quantitative reverse transcription polymerase chain reaction (RT-qPCR) across patient categories. Key findings include the elevated expression of CXCL1, CXCL2, IL1B, IL6, CXCL8 (IL8), PTGS2, and SPP1 in CRC tissues. Additionally, CXCL1, CXCL2, IL6, CXCL8, and PTGS2 showed significant expression changes in adenomatous polyps, suggesting their early involvement in carcinogenesis. This study uncovers a distinctive immunological signature in colorectal neoplasia among Norwegians, highlighting CXCL1, CXCL2, IL1B, IL6, CXCL8, PTGS2, and SPP1 as potential CRC biomarkers. These findings warrant further research to confirm their role and explore their utility in non-invasive screening strategies.

Response of soil microbial glycoside hydrolase family 6 cellulolytic population to lignocellulosic biochar reveals biochar stability toward microbial degradation.

Biochar produced from lignocellulosic biomass offers an opportunity to recycle waste into a valuable soil amendment. The application of biochar has been proposed to mitigate climate change by sequestering carbon in the soil. However, the field impact of biochar treatment on the cellulolytic microbial populations involved in the earlier steps of cellulose degradation is poorly understood. A field trial spanning three consecutive crop cycles of Zea mays was conducted in a degraded tropical Ultisol of Peninsular Malaysia. The soil was amended with two contrasting biochar made from oil palm kernel shells (pyrolyzed at 400°C) and rice husks (gasified at 800°C) with or without fertilizer supplementation. Soil samples were taken at each harvesting stage and analyzed for total organic carbon, labile active organic carbon, total cellulase, and β-glucosidase. Microbial glycoside hydrolase family 6 (GH6) cellulase genes and transcripts, involved in the early steps of cellulose degradation, were quantified from the extracted soil deoxyribonucleic acid (DNA) and ribonucleic acid (RNA), respectively. Total organic carbon, labile active organic carbon, and β-glucosidase activity were significantly increased, while no effect on total cellulase activity was found. Both biochars stimulated the total population (DNA-derived) abundance of soil microorganisms harboring the GH6 cellulase genes. The biochar amendment did not affect the active population (RNA-derived) of the GH6 cellulolytic community, showing no significant changes in transcript expression. This indirectly corroborates the role of biochar as a potential carbon sequester in the soil.

Crebanine, an aporphine alkaloid, induces cancer cell apoptosis through PI3K-Akt pathway in glioblastoma multiforme.

Glioblastoma multiforme (GBM) is one of the most prevalent and lethal primary central nervous system malignancies. GBM is notorious for its high rates of recurrence and therapy resistance and the PI3K/Akt pathway plays a pivotal role in its malignant behavior. Crebanine (CB), an alkaloid capable of penetrating the blood-brain barrier (BBB), has been shown to have inhibitory effects on proinflammatory molecules and multiple cancer cell lines via pathways such as PI3K/Akt. This study aims to investigate the efficacy and mechanisms of CB treatment on GBM. It is the first study to elucidate the anti-tumor role of CB in GBM, providing new possibilities for GBM therapy. Through a series of experiments, we demonstrate the significant anti-survival, anti-clonogenicity, and proapoptotic effects of CB treatment on GBM cell lines. Next-generation sequencing (NGS) is also conducted and provides a complete list of significant changes in gene expression after treatment, including genes related to apoptosis, the cell cycle, FoxO, and autophagy. The subsequent protein expressions of the upregulation of apoptosis and downregulation of PI3K/Akt are further proved. The clinical applicability of CB to GBM treatment could be high for its BBB-penetrating feature, significant induction of apoptosis, and blockage of the PI3K/Akt pathway. Future research is needed using in vivo experiments and other therapeutic pathways shown in NGS for further clinical or in vivo studies.

Identification of a highly drought-resistant pp7l hda6 mutant.

Plants have developed efficient strategies to counteract drought stress, including stomata closure, significant changes in nuclear gene expression, and epigenetic mechanisms. Previously, we identified Arabidopsis thaliana PROTEIN PHOSPHATASE7-LIKE (PP7L) as an extrachloroplastic protein that promotes chloroplast development. In addition, it was shown that PP7L is involved in high light and salt tolerance. Here, we demonstrate that the pp7l mutant can withstand prolonged periods of drought stress. Interestingly, despite impaired growth under standard growth conditions, photosynthetic efficiency recovers in pp7l mutant plants experiencing drought conditions. To assess the (post)transcriptional changes occurring in the pp7l mutant under different durations of drought exposure, we used an RNA-sequencing technique that allows the simultaneous detection of organellar and nuclear transcripts. Compared with the previously reported drought-responsive changes in the wild type, the drought-responsive changes in organellar and nuclear transcripts detected in the pp7l mutant were negligible. Our analysis of the data generated in this study and review and analysis of previous literature motivated us to create a pp7l hda6 (histone deacetylase 6) mutant, which exhibits remarkable drought resistance. Notably, the growth penalty associated with pp7l was alleviated in the double mutant, ruling out a dwarf effect on the drought-tolerant trait of this genotype. Future studies may consider that multiple loci and factors are involved in stress resistance and explore combinations of these factors to create even more resilient plants.

Dysregulated Transcript Expression but Not Function of the Glutamate Transporter EAAT2 in the Dorsolateral Prefrontal Cortex in Schizophrenia.

Schizophrenia (SCZ) is a serious mental illness with complex pathology, including abnormalities in the glutamate system. Glutamate is rapidly removed from the synapse by excitatory amino acid transporters (EAATs). Changes in the expression and localization of the primary glutamate transporter EAAT2 are found in the brain in central nervous system (CNS) disorders including SCZ. We hypothesize that neuronal expression and function of EAAT2 are increased in the frontal cortex in subjects diagnosed with SCZ. EAAT2 protein expression and glutamate transporter function were assayed in synaptosome preparations from the dorsolateral prefrontal cortex (DLPFC) of SCZ subjects and age- and sex-matched nonpsychiatrically ill controls. EAAT2 splice variant transcript expression was assayed in enriched populations of neurons and astrocytes from the DLPFC. Pathway analysis of publicly available transcriptomic datasets was carried out to identify biological changes associated with EAAT2 perturbation in different cell types. We found no significant changes in EAAT2 protein expression or glutamate uptake in the DLPFC in SCZ subjects compared with controls (n = 10/group). Transcript expression of EAAT2 and signaling molecules associated with EAAT2b trafficking (CaMKIIa and DLG1) were significantly altered in enriched populations of astrocytes and pyramidal neurons (P < .05) in SCZ (n = 16/group). These changes were not associated with antipsychotic medications. Pathway analysis also identified cell-type-specific enrichment of biological pathways associated with perturbation of astrocyte (immune pathways) and neuronal (metabolic pathways) EAAT2 expression. Overall, these data support the growing body of evidence for the role of dysregulation of the glutamate system in the pathophysiology of SCZ.

The Effect of Foliar Application with Naphthalene Acetic Acid and Potassium Nitrate on the Growth, Sex Ratio, and Productivity of Cucumbers (Cucumis sativas L.) under High Temperatures in Semi-Arid Areas

High temperatures in late spring, summer, and autumn are one of the main elements limiting cucumber production. Heat stress in cucumber plants leads to significant changes in sex expression and negative effects at the physiological stage, including decreased absorption of water and nutrients, decreased photosynthetic functions, and increased respiration, which in turn affects growth and production. Therefore, it is very difficult for cucumber plants exposed to heat stress to improve their productivity, especially in arid and semi-arid regions. Cucumber plants bear a variety of flower forms, including staminate, pistillate, and hermaphrodites that occur in different arrangements. In addition, maleness is considered the main problem in cucumber production, as it significantly reduces fruit yield. Recently, the growth, flowering, and productivity of plants have been controlled with the help of growth-regulating substances, and one of these substances is naphthalene acetic acid (NAA), which is of great importance in modifying sex in cucumber and various cucurbit crops. Two experiments were conducted during 2022 and 2023 during the summer season to study the effect of foliar spraying of two levels of potassium nitrate, 500 mg/L and 1000 mg/L, and two levels of NAA, 50 mg/L and 100 mg/L, individually and together, and the comparison treatment in nine treatments on growth traits, sex ratio, fruit traits and yield of cucumber (Cucumis sativas L.) plants in three replicates in a completely randomized block design. The results indicated that applying the treatment containing 1000 mg/L KNO3 + 100 mg/L NAA led to obtaining the highest value in most vegetable growth, flowering, and production parameters, with a significant increase compared to the control and all other treatments. Followed by treatments consisting of potassium nitrate and NAA, which gave high values in the traits of sex ratio %, fruit setting %, fruit yield, and most other traits. This study showed the importance of spraying with both NAA and potassium nitrate, individually or in combination, and the high combined effect of both in the treatments that contained each of them in the vegetative growth traits and floral traits such as the number of female flowers, the sexual ratio, fruit set percentage, weight fruit, length fruit, and the fruit yield in cucumber plants grown in the summer season with high temperatures

Tumor-matched and unmatched cancer associated fibroblasts exhibit differential effect on proliferation and FMOD and MMP9 gene expression in head and neck squamous cell carcinoma cells when cocultured in spheroids

BackgroundCancer-associated fibroblasts (CAFs) are the major cellular component of the tumor microenvironment and are known to affect tumor growth and response to various treatments. This study was undertaken to investigate the crosstalk between tumor-matched or unmatched CAFs and head and neck squamous cell carcinoma (HNSCC) cells regarding tumor growth and treatment response.MethodsThree HNSCC cell lines (LK0412, LK0902 and LK0923), were cocultured in 2D or in 3D with their tumor-matched CAFs, site matched CAFs from other tumors or normal oral fibroblasts (NOFs). Cell proliferation was assessed as the amount of Ki67 positive cells/ spheroid area in formalin-fixed- paraffin-embedded 3D spheroids stained with Ki67 antibody. Viability after seven days of cisplatin treatment was measured with CellTiter-Glo 3D Viability Assay. The mRNA expression of CAF-associated markers (ACTA2, COL1A2, FAP, PDGFRα, PDGFRβ, PDPN, POSTN and S100A4) in CAFs before and after coculture with tumor cells as well as mRNA expression of CAF-induced genes (MMP1, MMP9 and FMOD) in tumor cells separated from CAFs after co-culture was measured with RT-qPCR. The expression of selected protein biomarkers was validated with immunohistochemistry based on previous mRNA expression results.ResultsThe proliferation of the LK0412 and LK0902 tumor spheroids varied significantly when cocultured with different CAFs and NOFs as shown by Ki-67 positive cells. RT‒qPCR analysis revealed different molecular profile of the analyzed HNSCC-derived CAFs concerning the expression of CAF-associated markers. The interaction between CAFs and HNSCC cells was more pronounced after coculture with unmatched CAFs as shown by changes in mRNA expression pattern of CAF-specific markers. Additionally, the unmatched CAFs significantly upregulated the mRNA expression of MMP1, MMP9 and FMOD in tumor cells compared to tumor-matched CAFs.ConclusionOur results indicate that tumor-matched CAFs are unique for each tumor and affect the proliferation and the gene/protein expression of tumor cells in a distinct manner. The interaction between tumor unmatched CAFs and HNSCC cells in the tumor spheroids is associated with significant changes in the mRNA expression of CAF-specific markers and significant increases in FMOD and MMP9 in tumor cells compared to when cocultured with tumor-matched CAFs. Taken together, our results show how important the selection of CAFs is to get a reliable in vitro model that mimics the patients’ tumor.

Inflammasome activation dynamics within the first 24 hours in patients treated with PCI for myocardial infarction is influenced by the age of patients, preliminary report

Abstract Funding Acknowledgements Type of funding sources: Public Institution(s). Main funding source(s): Presented study was financially supported by the Mini-student Grant for medical students of Medical University of Warsaw, No 24/M/MG/N/21 and Grant for Young Researchers of The Medical University of Warsaw, No 2W6/1/M/MBS/N/21. Background Until today, majority of data on the possible role of inflammasome activation in acute coronary syndrome (ACS) patients comes from preclinical studies. Purpose We aimed to determine the dynamics of the expression profile of selected inflammasome genes and inflammasome related factors by quantitative real-time polymerase chain reaction (qRT-PCR) in peripheral blood mononuclear cells (PBMCs) of patients in the acute phase of ACS. Methods In the study group the blood samples for RT-PCR were collected 3 times: at the beginning of the percutaneous coronary intervention (PCI) for ACS, 3h, and 24h after PCI. The study group samples were compared with blood collected from 10 patients with no significant coronary lesions identified during coronary angiography. The relative expression of evaluated genes was analyzed in relation to the β-actin gene in relation to the expression in 0-hour sample using the ΔΔCT method. Based on the literature, we defined a significant change in gene expression at a fold change of 1.5. Results For this preliminary study the population of 19 patients aged 64±13and 84% presented with ST-elevation myocardial infarction (STEMI). Expression of evaluated genes at 3h and 24h relative to 0h sample is presented in picture 1. Gene expression did not differ significantly between the study and control group except for NLR Family Pyrin Domain Containing 6 (NLRP6) (at 3h post-PCI: ΔΔCT=2.5425 vs 0.0005; p=0.002, 24h post-PCI: ΔΔCT=2.1794 vs 0.0005; p=0.009). However, we observed significant upregulation in samples taken at 3h post-PCI relative to 0h level for AIM2 [fold change relative to 0h 1.52 (0.63; 4.67)], IL-1β [2.81 (1.59; 5.93)], IL-18 [2.07 (0.59; 10.14)] and at 24 h for AIM2 [1.90 (0.78; 2.52)], IL-1β [5.16 (2.23; 13.32)] and IL-18 [2.20 (0.59; 5.96)] in the younger subgroup of patients (≤the median age of the study population which was equal 66 years old). We performed a comparison of younger (≤66) vs. older (&amp;gt;66) patients at 3h and 24h post-PCI and observed significantly higher relative gene expression in the younger subgroup compared to the older subgroup of patients at 3h in case of IL-1β (p=0.03), and at 24h in case of NLR Family Pyrin Domain Containing 1(NLRP1)(p=0.016), AIM2 (p=0.04), NLR Family Pyrin Domain Containing 12(NRLP12)(p=0.06), IL-1 (p=0.04), and IL-18 (p=0.03) – the exact fold change values are presented in Table 1. Conclusion NLRP6 is significantly upregulated in ACS patients compared to control subjects in the first 24h post-PCI. We also observed significant dynamics in NLRP3 inflammasome related factors, including IL-1β, IL-18, in the first 24h post-PCI for MI. Also, the activation of the inflammasome genes (NLRP1, NLRP12) and inflammasome effectors may present different dynamics dependent on the age of the patients, and it might be helpful in careful selection of the timepoint for the future administration of inflammasome dedicated inhibitors.Relative genes expressionFold change in gene expression

Immune cell populations in the tumour environment following calcium electropora-tion for cutaneous metastasis: a histopathological study.

Calcium electroporation (CaEP) involves injecting calcium into tumour tissues and using electrical pulses to create membrane pores that induce cell death. This study assesses resultant immune responses and histopathological changes in patients with cutaneous metastases. The aimed cohort comprised 24 patients with metastases exceeding 5 mm. Tumours were treated once with CaEP (day 0) or twice (day 28). Biopsies were performed on days 0 and 2, with additional samples on days 7, 28, 30, 35, 60, and 90 if multiple tumours were treated. The primary endpoint was the change in tumour-infiltrating lymphocytes (TILs) two days post-treatment, with secondary endpoints evaluating local and systemic immune responses via histopathological analysis of immune markers, necrosis, and inflammation. Seventeen patients, with metastases primarily from breast cancer (14 patients), but also lung cancer (1), melanoma (1), and urothelial cancer (1), completed the study. Of the 49 lesions treated, no significant changes in TIL count or PD-L1 expression were observed. However, there was substantial necrosis and a decrease in FOXP3-expression (p = 0.0025) noted, with a slight increase in CD4+ cells but no changes in CD3, CD8, or CD20 expressions. Notably, four patients showed reduced tumour invasiveness, including one case of an abscopal response. This exploratory study indicates that CaEP can be an effective anti-tumour therapy potentially enhancing immunity. Significant necrosis and decreased regulatory lymphocytes were observed, although TIL count remained unchanged. Several patients exhibited clinical signs of immune response following treatment.

Pathological Changes and Metabolic Adaptation in the Myocardium of Rats in Response to Chronic Variable Mild Stress.

Chronic variable mild stress (CVS) in rats is a well-established paradigm for inducing depressive-like behaviors and has been utilized extensively to explore potential therapeutic interventions for depression. While the behavioral and neurobiological effects of CVS have been extensively studied, its impact on myocardial function remains largely unexplored. To induce the CVS model, rats were exposed to various stressors over 40 days. Behavioral assessments confirmed depressive-like behavior. Biochemical analyses revealed alterations in myocardial metabolism, including changes in NAD+ and NADP+, and NADPH concentrations. Free amino acid analysis indicated disturbances in myocardial amino acid metabolism. Evaluation of oxidative DNA damage demonstrated an increased number of abasic sites in the DNA of rats exposed to CVS. Molecular analysis showed significant changes in gene expression associated with glucose metabolism, oxidative stress, and cardiac remodeling pathways. Histological staining revealed minor morphological changes in the myocardium of CVS-exposed rats, including increased acidophilicity of cells, collagen deposition surrounding blood vessels, and glycogen accumulation. This study provides novel insights into the impact of chronic stress on myocardial function and metabolism, highlighting potential mechanisms linking depression and cardiovascular diseases. Understanding these mechanisms may aid in the development of targeted therapeutic strategies to mitigate the adverse cardiovascular effects of depression.