The elusive male microbiome: revealing the link between the genital microbiota and fertility. Critical review and future perspectives.

There is a growing focus on understanding the role of the male microbiome in fertility issues. Although research on the bacterial communities within the male reproductive system is in its initial phases, recent discoveries highlight notable variations in the microbiome's composition and abundance across distinct anatomical regions like the skin, foreskin, urethra, and coronary sulcus. To assess the relationship between male genitourinary microbiome and reproduction, we queried various databases, including MEDLINE (available via PubMed), SCOPUS, and Web of Science to obtain evidence-based data. The literature search was conducted using the following terms "gut/intestines microbiome," "genitourinary system microbiome," "microbiome and female/male infertility," "external genital tract microbiome," "internal genital tract microbiome," and "semen microbiome." Fifty-one relevant papers were analyzed, and eleven were strictly semen quality or male fertility related. The male microbiome, especially in the accessory glands like the prostate, seminal vesicles, and bulbourethral glands, has garnered significant interest because of its potential link to male fertility and reproduction. Studies have also found differences in bacterial diversity present in the testicular tissue of normozoospermic men compared to azoospermic suggesting a possible role of bacterial dysbiosis and reproduction. Correlation between the bacterial taxa in the genital microbiota of sexual partners has also been found, and sexual activity can influence the composition of the urogenital microbiota. Exploring the microbial world within the male reproductive system and its influence on fertility opens doors to developing ways to prevent, diagnose, and treat infertility. The present work emphasizes the importance of using consistent methods, conducting long-term studies, and deepening our understanding of how the reproductive tract microbiome works. This helps make research comparable, pinpoint potential interventions, and smoothly apply microbiome insights to real-world clinical practices.

The effect of local anaesthetics on apoptosis and NETosis of human neutrophils in vitro: comparison between lidocaine and ropivacaine.

There are immunological consequences to the method by which neutrophils undergo cell death. Neutrophil apoptosis, called silent death, leads to the resolution of inflammation, while NETosis deepens and prolongs the inflammatory response and is associated with a worse prognosis of severe infections, e.g., sepsis. Besides nociceptive inhibition, local anaesthetics modulate leukocyte functions, even at low, clinically relevant concentrations. There is currently no data on ropivacaine NETosis, and this study aimed to evaluate the impact of clinical concentrations of ropivacaine (0.0007, 0.007 and 1.4 mmol/L) and lidocaine (0.002, 0.02 and 4 mmol/L) on apoptosis and NETosis of adult peripheral blood neutrophils after 2 h of incubation. Neutrophil identification, apoptosis and NETosis were evaluated by flow cytometry using forward and side scatter characteristics and fluorescent labelling: CD15 for neutrophils identification; Annexin V and propidium iodide for apoptosis and citrullinated histone H3 and myeloperoxidase for NETosis. Lidocaine (4 mmol/L) and ropivacaine (1.4 mmol/L) induced early apoptosis in resting but not in stimulated neutrophils. Low doses of ropivacaine (0.0007 and 0.007 mmol/L) decreased the number of late apoptotic neutrophils, and the lowest dose slightly increased their viability. None of the drugs induced NETosis in resting neutrophils but decreased NETosis at clinical concentrations compared to PMA-stimulated 4 mM lidocaine, PMA-stimulated control, and 1.4 mM ropivacaine. The effect of lidocaine and ropivacaine on apoptosis and NETosis depended on neutrophil stimulation and drug concentrations. Ropivacaine tends to be cytoprotective at concentrations observed in plasma under local anaesthesia. Lidocaine enhanced NETosis at high concentration only in stimulated neutrophils. Thus, both drugs have the ability to change the course of inflammation.

PELP1 and SRC kinase as important molecules in the estrogen-mediated pathway in human testis and epididymis.

BACKGROUND: Estrogens have pleiotropic mechanisms of action, and their cellular transduction pathways can modulate various proteins with differential tissue expression. Proline-, glutamic acid-, and leucine-rich protein 1 (PELP1) is one such protein whose role seems important, although little is known about this protein. However, very little is known about the expression of modulators involved in the estrogen-mediated pathways in the tissues of the male reproductive tract. METHODS: In this study, we obtained autopsy specimens of testis and epididymis from 13 men of Caucasian descent. Expression levels were analyzed for both estrogen receptors (ESR1 and ESR2) and their co-regulators, including PELP1 and kinase c-Src (SRC). RESULTS: Protein expression was confirmed with western blot and immunocytochemistry techniques. The expression of both SRC and PELP1 was significantly higher in the testis compared to the epididymis (p=0.040 and p=0.002, respectively). Furthermore, a significant, positive correlation was observed between SRC and PELP1, regardless of tissue type p<0.0001, R=0.78). In the testis, PELP1 expression positively correlated with ESR1 expression (p=0.367, R=0.6). CONCLUSIONS: Our study suggests a possible relationship between PELP1, SRC, and ESR1 in the human testis and epididymis. This study makes a valuable contribution to the field of estrogen-mediated pathways in the male reproductive tract and describes trends of analyzed genes' expression and presence. We think our results may open some new research directions of the estrogen signaling in the male reproductive system.

MicroRNA expression profile analysis in human skeletal muscle tissue: Selection of critical reference.

MicroRNAs (miRNAs) are highly conserved small non-coding RNAs, that modulate gene expression by targeting messenger RNA of many processes. Thus, miRNAs are key regulators of both physiological and pathological settings. Reliable results of quantitative miRNA evaluation depend on suitable reference genes (RGs) for data normalization. To date, no consensus has been reached on the best RG for muscle tissue. We assessed RGs stability in skeletal muscle tissue in patients with spinal deformity. Ninety tissue samples were obtained from the deep paravertebral muscles from the convex and concave sides of the spinal curvature, as well as the superficial paraspinal muscles. We evaluated the stability of twelve miRNAs (hsa-miR-1-3p, hsa-miR-1-5p, hsa-miR-26b-5p, hsa-miR-92a-3p, hsa-miR-133a-3p, hsa-miR-133a-5p, hsa-miR-133b, hsa-miR-191-5p, hsa-miR-206, hsa-miR-208b-5p, hsa-miR-486-5p, hsa-miR-499a-5p), finding three to be indicative of reference miRNA, and nine as muscle-tissue specific. Stability was quantified using four statistical tools and a comprehensive ranking system. Three miRNAs were indicated as the most stable, and we assessed hsa-miR-486-5p as the most, and hsa-miR-208b-5p as the least suitable RGs for miRNA quantitative analyses. We recommend using a minimum of three RGs miRNA to normalize RT-qPCR data. Finally, qPCR efficiency should always be considered. To obtain consistent results, data normalization in muscle tissue is required.

Clinical Implications of mTOR Expression in Papillary Thyroid Cancer-A Systematic Review.

Papillary thyroid cancer (PTC) comprises approximately 80% of all thyroid malignancies. Although several etiological factors, such as age, gender, and irradiation, are already known to be involved in the development of PTC, the genetics of cancerogenesis remain undetermined. The mTOR pathway regulates several cellular processes that are critical for tumorigenesis. Activated mTOR is involved in the development and progression of PTC. Therefore, we performed a systematic review of papers studying the expression of the mTOR gene and protein and its relationship with PTC risk and clinical outcome. A systematic literature search was performed using PubMed, Embase, and Scopus databases (the search date was 2012-2022). Studies investigating the expression of mTOR in the peripheral blood or tissue of patients with PTC were deemed eligible for inclusion. Seven of the 286 screened studies met the inclusion criteria for mTOR gene expression and four for mTOR protein expression. We also analyzed the data on mTOR protein expression in PTC. We analyzed the association of mTOR expression with papillary thyroid cancer clinicopathological features, such as the TNM stage, BRAF V600E mutation, sex distribution, lymph node and distant metastases, and survival prognosis. Understanding specific factors involved in PTC tumorigenesis provides opportunities for targeted therapies. We also reviewed the possible new targeted therapies and the use of mTOR inhibitors in PTC. This topic requires further research with novel techniques to translate the achieved results to clinical application.

Expression of estrogen receptors, PELP1, and SRC in human spermatozoa and their associations with semen quality.

Sperm cells are target cells for both estrogens and xenoestrogens. Due to the specific structure of spermatozoa, these hormonal compounds may act on sperm in a non-genomic mechanism only. However, the ESR-mediated signaling pathways are still poorly understood. In this study, we obtained 119 samples from male participants of Caucasian descent who donated semen for standard analysis. We analyzed gene expression of estrogen receptors (ESR1 and ESR2) and their coregulators-proline-, glutamic acid-, and leucine-rich protein 1 (PELP1), and cellular kinase c-Src (SRC). RNA level was established using reverse-transcribed RNA as a template, followed by a polymerase chain reaction. Proteins' presence was confirmed by western blot and immunocytochemistry techniques. "Normal" values of semen parameters were defined as follows: > 32% sperm with progressive motility, > 4% sperm cells with normal morphology, > 15 × 106 sperm per mL, > 58% live spermatozoa and leukocyte amount < 106 cells per mL, according to WHO 2010 reference. Semen parameters that deviated from these "normal" values were labeled as "abnormal". Gene expression ratios revealed significant, moderate, and negative correlations for ESR1/ESR2 and weak, negative ESR2/PELP1 correlations in the subgroup of patients with abnormal values of semen parameters. In addition, SRC/PELP1 was moderately and positively correlated in the subgroup with parameters within the reference values established by WHO 2010. Our study showed that both PELP1 scaffolding protein and SRC kinase might influence semen quality via ESRs. It seems that not the expression of a single gene may affect the sperm quality, but more gene-to-gene mutual ratio. Characterization of estrogen-signaling pathway-related genes' modulated expression in sperm cells could aid in better understanding sperm biology and quality.

Ciliary phenotyping in renal epithelial cells in a cranioectodermal dysplasia patient with WDR35 variants.

Background: Cranioectodermal dysplasia (CED) is a skeletal autosomal recessive ciliopathy. The characteristic clinical features of CED are facial dysmorphisms, short limbs, narrow thorax, brachydactyly, ectodermal abnormalities, and renal insufficiency. Thus far, variants in six genes are known to be associated with this disorder: WDR35, IFT122, IFT140, IFT144, IFT52, and IFT43. Objective: The goal of this study was to perform cilium phenotyping in human urine-derived renal epithelial cells (hURECs) from a CED patient diagnosed with second-stage chronic kidney disease (CKD) and three unrelated and unaffected pediatric controls. Methods: Genetic analysis by WDR35 screening was performed in the affected individual. Cilium frequency and morphology, including cilium length, height, and width, were evaluated by immunofluorescence (IF) experiments in hURECs using two markers visualizing the ciliary axoneme (Acet-Tub and ARL13B) and the base of the cilium (PCNT). The IF results were analyzed using a confocal microscope and IMARIS software. Results: WDR35 analysis revealed the presence of a known nonsense p. (Leu641*) variant and a novel missense variant p. (Ala1027Thr). Moreover, comparative genomic hybridization analysis showed that the patient carries a microdeletion on chromosome 7q31.1. Ciliary phenotyping performed on hURECs showed morphological differences in the patient's cilia as compared to the three controls. The cilia of the CED patient were significantly wider and longer. Conclusion: The obtained results suggest that CED-related second-stage CKD might be associated with cilia abnormalities, as identified in renal epithelial cells from a CED patient harboring variants in WDR35. This study points out the added value of hURECs in functional testing for ciliopathies.

Impact of Biometric Patient Data, Probiotic Supplementation, and Selected Gut Microorganisms on Calprotectin, Zonulin, and sIgA Concentrations in the Stool of Adults Aged 18-74 Years.

Alterations to the intestinal barrier may be involved in the pathogenesis of various chronic diseases. The diagnosis of mucosal barrier disruption has become a new therapeutic target for disease prevention. The aim of this study was to determine whether various patient demographic and biometric data, often not included in diagnostic analyses, may affect calprotectin, zonulin, and sIgA biomarker values. Stool markers' levels in 160 samples were measured colorimetrically. The analysis of twenty key bacteria (15 genera and 5 species) was carried out on the basis of diagnostic tests, including cultures and molecular tests. The concentrations of selected markers were within reference ranges for most patients. The sIgA level was significantly lower in participants declaring probiotics supplementation (p = 0.0464). We did not observe differences in gastrointestinal discomfort in participants. We found significant differences in the sIgA level between the 29-55 years and >55 years age-related intervals groups (p = 0.0191), together with a significant decreasing trend (p = 0.0337) in age-dependent sIgA concentration. We observed complex interdependencies and relationships between their microbiota and the analyzed biomarkers. For correct clinical application, standardized values of calprotectin and sIgA should be determined, especially in elderly patients. We observed a correlation between the composition of the gut community and biomarker levels, although it requires further in-depth analysis.

Differential Expression of HIF1A, EPAS1, and VEGF Genes in Benign and Malignant Ovarian Neoplasia.

Ovarian cancer (OC) has the highest mortality rate of all gynecological malignancies. Moreover, at the time of the first clinical manifestation, most patients have an advanced stage of the disease. Our study examined differences in mRNA levels of hypoxia-inducible factor 1-alpha (HIF1A); endothelial PAS domain protein 1, also known as hypoxia-inducible factor 2-alpha (HIF2A/EPAS1); and vascular endothelial growth factor A (VEGFA) between cancerous tissue, benign hyperplastic changes in the ovary, and normal tissue. Our cohorts consisted of 52 patients diagnosed with OC (n = 55), benign non-cancerous changes (n = 21), and normal tissue samples (n = 38). The mRNA expression level was evaluated using RT-qPCR. We found that gene expression changes were visible not only in the case-control study, but also along with changes in severity. Additionally, the gene expression was differentiated in age, BMI, menopausal status, and the number of comorbidy-related groups. Furthermore, our findings demonstrate that analyzing the correlation between genes is essential. In a case-to-case and case-to-control study, we observed disturbances in the expression levels of interdependent genes. Our findings suggest that mutual association in the expression of both HIF1A and HIF2A/EPAS1 with VEGFA has prognostic importance for patients with OC. Our observations may help identify patients for clinical trials aimed at inhibiting the hypoxia-induced neovascularization-dependent pathways.

Association of LBX1 Gene Methylation Level with Disease Severity in Patients with Idiopathic Scoliosis: Study on Deep Paravertebral Muscles.

Idiopathic scoliosis (IS) is a multifactorial disease with a genetic background. The association of Ladybird Homeobox 1 (LBX1) polymorphisms with IS has been proven in multiple studies. However, the epigenetic mechanisms have not been evaluated. This study aimed to evaluate the LBX1 methylation level in deep paravertebral muscles in order to analyze its association with IS occurrence and/or IS severity. Fifty-seven IS patients and twenty non-IS patients were examined for the paravertebral muscles' methylation level of the LBX1 promoter region. There was no significant difference in methylation level within paravertebral muscles between patients vs. controls, except for one CpG site. The comparison of the paravertebral muscles' LBX1 promoter region methylation level between patients with a major curve angle of ≤70° vs. >70° revealed significantly higher methylation levels in 17 of 23 analyzed CpG sequences at the convex side of the curvature in patients with a major curve angle of >70° for the reverse strand promoter region. The association between LBX1 promoter methylation and IS severity was demonstrated. In patients with severe IS, the deep paravertebral muscles show an asymmetric LBX1 promoter region methylation level, higher at the convex scoliosis side, which reveals the role of locally acting factors in IS progression.

Estrogen Receptor Type 1 and Type 2 Presence in Paravertebral Skeletal Muscles: Expression Level and Relation to Phenotype in Children with Idiopathic Scoliosis.

The study aimed to detect the presence and assess the expression levels of the estrogen receptors type 1 (ESR1) and type 2 (ESR2) within paravertebral skeletal muscles of female patients with idiopathic scoliosis (IS) in relation to phenotype parameters. Intraoperatively, the muscle samples were obtained from 35 adolescent females. The RT-qPCR, western blot and immunohistochemistry techniques were applied. The ESR1 and ESR2 were detected within paravertebral skeletal muscle cells, either the superficial or the deep ones. The ESR1 expression level was significantly higher in the deep muscles compared to the superficial ones. A left-right asymmetry of the ESR1 and ESR2 expression level was demonstrated in the deep muscles. There was a significant relationship between the expression asymmetry and either the Cobb angle or the progression risk factor: both parameters decreased to the smallest values in the case of symmetric ESR1 or ESR2 expression, while they increased with increasing expression asymmetry. In conclusion, the ESR1 and ESR2 presence was confirmed in skeletal paravertebral muscles of patients with idiopathic scoliosis. The increased expression level and asymmetry of estrogen receptors in deep skeletal muscles was related to increasing scoliotic deformity magnitude or increasing risk of deformity deterioration. These findings may highlight the etiopathogenesis of IS in children.

Commercial microbiota test revealed differences in the composition of intestinal microorganisms between children with autism spectrum disorders and neurotypical peers.

The early-life modifications of intestinal microbiota may impact children's subsequent emotional and cognitive development. Studies show that some bacteria species in gut microbiota, and the lack of others, may play a key role in autism spectrum disorders (ASD) development. Fecal samples were obtained from three groups of children: 16 healthy, 24 with allergies (ALG), and 33 with ASD (probiotics and non-probiotics users). The analysis was carried out according to the KyberKompakt Pro protocol. We observed a significantly higher level of Klebsiella spp. in the healthy children from the non-probiotics group, considering three groups. In the same group, Bifidobacterium spp. the level was lower in ASD compared to neurotypical individuals. In healthy children who did not use probiotics, strong positive correlations were observed in E. coli and Enterococcus spp. and Bacteroides and Klebsiella spp., and a negative correlation for Akkermansia muciniphila with both Klebsiella spp. and Bacteroides spp. In the ASD group who take probiotics, a strongly negative correlation was observed in Lactobacillus spp., and both Faecalibacterium prausnitzii and Akkermansia muciniphila levels. In the ALG group, the strongest, negative correlation was found between Enterococcus spp. and Lactobacillus spp. as in Akkermansia muciniphila and Bifidobacterium spp. The simple commercial test revealed minor differences in the composition of intestinal microorganisms between children with autism spectrum disorders and neurotypical peers.

Phoenixin as a New Target in the Development of Strategies for Endometriosis Diagnosis and Treatment.

Small integral membrane protein 20/phoenixin (SMIM20/PNX) and its receptor GPR173 (G Protein-Coupled Receptor 173) play a role in the regulation of the hypothalamic-pituitary-gonadal axis (HPG). The aim of the study was to determine PNX, FSH, LH, and 17ß-estradiol association in women with endometriosis, and the expression of SMIM20/PNX signaling via GPR173. Serum PNX, FSH, LH, and 17ß-estradiol concentrations were measured by enzyme and electrochemiluminescence immunoassay. SMIM20/PNX and GPR173 expression in the eutopic and ectopic endometrium was assessed by qPCR and immunohistochemistry. Reduced PNX level, increased LH/FSH ratio and elevated 17ß-estradiol concentration were found in patients with endometriosis. No differences in SMIM20 expression were observed between the studied endometria. GPR173 expression was lower in ectopic than in eutopic endometria. SMIM20 expression was mainly restricted to stroma. GPR173 was detected in some eutopic and ectopic stromal cells and in eutopic glandular epithelial cells. Discriminant analysis indicates the diagnostic relevance of PNX and LH/FSH ratio in patients with endometriosis. In women with endometriosis, reduced PNX levels and GPR173 expression may be responsible for HPG axis dysregulation. These new insights may contribute to a better understanding of the pathophysiology of endometriosis and provide the basis for a new strategy for diagnosis and treatment of endometriosis.

Altered Expression of ESR1, ESR2, PELP1 and c-SRC Genes Is Associated with Ovarian Cancer Manifestation.

Ovarian cancer remains the leading cause of death due to gynecologic malignancy. Estrogen-related pathways genes, such as estrogen receptors (ESR1 and ESR2) and their coregulators, proline-, glutamic acid-, and leucine-rich protein 1 (PELP1), and proto-oncogene tyrosine-protein kinase c-Src (SRC) are involved in ovarian cancer induction and development, still they require in-depth study. In our study, tissue samples were obtained from 52 females of Caucasian descent (control group without cancerous evidence (n = 27), including noncancerous benign changes (n = 15), and the ovarian carcinoma (n = 25)). Using quantitative analyses, we investigated ESRs, PELP1, and SRC mRNA expression association with ovarian tumorigenesis. Proteins' presence and their location were determined by Western blot and immunohistochemistry. Results showed that PELP1 and SRC expression levels were found to differ in tissues of different sample types. The expression patterns were complex and differed in the case of ovarian cancer patients compared to controls. The most robust protein immunoreactivity was observed for PELP1 and the weakest for ESR1. The expression patterns of analyzed genes represent a potentially interesting target in ovarian cancer biology, especially PELP1. This study suggests that specific estrogen-mediated functions in the ovary and ovary-derived cancer might result from different local interactions of estrogen with their receptors and coregulators.

Methylation of Estrogen Receptor 1 Gene in the Paraspinal Muscles of Girls with Idiopathic Scoliosis and Its Association with Disease Severity.

Idiopathic scoliosis (IS) is a multifactorial disease with epigenetic modifications. Tissue dependent and differentially methylated regions (T-DMRs) may regulate tissue-specific expression of the estrogen receptor 1 gene (ESR1). This study aimed to analyze methylation levels within T-DMR1 and T-DMR2 and its concatenation with ESR1 expression of IS patients. The study involved 87 tissue samples (deep paravertebral muscles, both on the convex and the concave side of the curve, and from back superficial muscles) from 29 girls who underwent an operation due to IS. Patient subgroups were analyzed according to Cobb angle ≤70° vs. >70°. Methylation was significantly higher in the superficial muscles than in deep paravertebral muscles in half of the T-DMR1 CpGs and all T-DMR2 CpGs. The methylation level correlated with ESR1 expression level on the concave, but not convex, side of the curvature in a majority of the T-DMR2 CpGs. The T-DMR2 methylation level in the deep paravertebral muscles on the curvature's concave side was significantly lower in patients with a Cobb angle ≤70° in four CpGs. DNA methylation of the T-DMRs is specific to muscle tissue location and may be related to ESR1 expression regulation. Additionally, the difference in T-DMR2 methylation may be associated with IS severity.

Negative psychological aspects of working with experimental animals in scientific research.

The aim of the study was to reveal the negative psychological aspects of using animals by scientists and to determine whether the emotional tensions and stress are associated with performing experiments on animals. All 150 participants of the study conduct experiments on animals in their work. Computer-assisted web interviewing, was used to collect the data. Correlation matrices for factorial analysis of main component loads and cluster analysis have been calculated as grouping methods revealed two different categories of researchers, which were mostly distinguished by acceptance and aversion to animal testing and animal welfare. The main findings demonstrated, that there is a group of respondents who feel discomfort when performing experiments on animals. Especially young people involved in animal testing, feel remorse, emotional tension and helplessness.

EZH2 and SMYD3 expression in papillary thyroid cancer.

Recent studies have revealed the significant role of SMYD3 and EZH2 genes in the development and aggressiveness of numerous types of malignant tumor. Therefore, the present study aimed to investigate the expression of SMYD3 and EZH2 in papillary thyroid cancer, and to determine the correlation between the expression of these genes and clinical characteristics. Resected thyroid tissue samples from 62 patients with papillary thyroid cancer were investigated. Thyroid tissue derived from the healthy regions of removed nodular goiters from 30 patients served as the control group. Reverse transcription-quantitative PCR analysis was employed to detect relative mRNA expression levels. Primer sequences and TaqMan® hydrolysis probe positions for EZH2 and SMYD3 were determined using the Roche Universal ProbeLibrary Assay Design Center version 2.50. EZH2 expression was detected in all thyroid cancer samples and in 83.3% of benign lesions. Notably, EZH2 was revealed to be upregulated in thyroid cancer tissues compared with control tissues (P=0.0002). EZH2 expression was positively correlated with tumor stage (P<0.0001; r=0.504), and multiple comparison analysis revealed that the highest expression of EZH2 was detected in samples staged pT4 (P=0.0001). SMYD3 expression was detected in all thyroid cancer samples and in 96.7% of healthy thyroid tissues; notably, the expression levels were similar in both groups. In addition, there was no correlation between SMYD3 expression and the aggressiveness of papillary thyroid cancer. In conclusion, overexpression of the EZH2 gene may be associated with the development of papillary thyroid cancer and EZH2 may be a potential therapeutic target in papillary thyroid cancer.

Methylation of estrogen receptor 2 (ESR2) in deep paravertebral muscles and its association with idiopathic scoliosis.

Idiopathic scoliosis (IS) is one of the most common spinal disorders in adolescents. Despite many studies, the etiopathogenesis of IS is still poorly understood. In recent years, the role of epigenetic factors in the etiopathogenesis of IS has been increasingly investigated. It has also been postulated that the development and progression of the disease is related to gender and puberty, and could be associated with estrogen action. Estrogen hormones act via estrogen receptor 1 (ESR1) and estrogen receptor 2 (ESR2). It has been suggested that ESR2 expression is dependent on methylation within its gene promoter. So far, no studies have evaluated local, tissue-specific DNA methylation in patients with IS. Thus, our study aimed to analyze the methylation and expression level of ESR2 in the paraspinal muscles of the convex and concave side of the IS curvature. The methylation level within ESR2 promoter 0N, but not exon 0N, was significantly higher on the concave side of the curvature compared to the convex side. There was no significant correlation between ESR2 expression and methylation level in the promoter 0N on the convexity of thoracic scoliosis, whereas, on the concave side of the curvature, we observed a moderate negative correlation. There was no difference in the methylation levels of the ESR2 promoter and exon 0N between groups of patients with Cobb angle ≤ 70° and > 70° on the concave and convex side of the curvature. We also found no statistically significant correlation between the Cobb angle value and the mean methylation level in either the ESR2 promoter or exon 0N on the convex or concave side of the curvature. Our findings demonstrate that DNA methylation at the ESR2 promoter in deep paravertebral muscle tissue is associated with the occurrence but not with the severity of idiopathic scoliosis.

Cerebral tissue oxygenation during cranial osteopathic CV4 procedure in newborns.

BACKGROUND: The cranial osteopathic manipulative medicine has been shown to alter regional cerebral tissue oxygenation (cStO2) in adult patients; however, there are no reports regarding the neonatal population. OBJECTIVES: To assess the influence of compression of the 4th ventricle (CV4) osteopathic procedure on cStO2 in neonates. MATERIAL AND METHODS: Thirty-one patients born between 25 and 39 weeks of gestation were screened for inclusion in the neonatal unit. Twenty-two infants presenting with hyperstimulation of autonomous nervous system (ANS) according to the Neonatal Behavioral Assessment Scale were enrolled in the study. Near-infrared spectroscopy was used for continuous cStO2 monitoring; pulse oximeter oxygen saturation (SpO2) and heart rate (HR) measured with pulse oximetry were simultaneously monitored 10 min before CV4, during the therapy and 10 min after it was stopped. RESULTS: Patients' condition remained stable throughout the study. There were no significant differences in the mean cStO2 values recorded before (69 ±8%), during (69 ±8%) and after CV4 (70 ±8%; p > 0.05). Mean SpO2 was almost constant during the study (96 ±4% before, 95 ±3% during and 95 ±4% after the intervention). Heart rate was also stable pre-, during and post-therapy (153 ±21 min, 151 ±18 min and 151 ±20/min, respectively). CONCLUSIONS: Compression of the 4th ventricle osteopathic procedure does not influence the cStO2 in newborns. This method seems to be well-tolerated but its clinical efficacy needs to be further investigated in this group of patients.

CHD7 gene polymorphisms in female patients with idiopathic scoliosis.

BACKGROUND: The CHD7 (chromosome domain helicase DNA binding protein 7) gene has been associated with familial idiopathic scoliosis (IS) in families of European descent. The CHD7 single-nucleotide polymorphisms have never been studied in Polish Caucasian IS patients. METHODS: The aim of this study was to investigate the relationship of CHD7 gene polymorphisms with susceptibility to or progression of IS in Polish Caucasian females. The study group comprised 211 females who underwent clinical, radiological and genetic examination. The study group was analyzed in three subgroups according to: (1) Cobb angle (Cobb angle ≤30° vs. Cobb angle ≥35°), (2) age of diagnosis (adolescent IS vs. early-onset IS) and (3) rate of progression (non-progressive vs. slowly progressive vs. rapidly progressive IS). The control group comprised 83 females with no scoliosis and with a negative family history who underwent clinical and genetic examination. In total six CHD7 gene polymorphisms were examined. Three polymorphisms (rs1017861, rs13248429, and rs4738813) were examined by RFLP (restriction fragment length polymorphism) analysis, and three were quantified by Sanger sequencing (rs78874766, rs4738824, and rs74797613). RESULTS: In rs13248429, rs78874766, and rs74797613 polymorphisms only the wild allele was present. The rs1017861 polymorphism demonstrated an association with IS susceptibility (p < 0.01). Two polymorphisms, rs1017861 and rs4738813, were associated with curve severity and progression rate (p < 0.05). None of the evaluated polymorphisms in CHD7 gene showed any association with the age of IS onset. CONCLUSIONS: The polymorphism rs1017861 in CHD7 gene showed an association with IS susceptibility. Two polymorphisms (rs1017861 and rs4738813) were associated with curve severity and progression rate. None of the evaluated polymorphisms in CHD7 gene showed any association with the age of IS onset. Further evaluation of CHD7 gene should be considered as IS modifying factor.