Association of OXTR, AVPR1a, LNPEP, and CD38 Genes' Expression with the Clinical Presentation of Autism Spectrum Disorder.

Autism spectrum disorder (ASD) is a complex neurodevelopmental disorder that affects social interactions, communication, and behavior. Although the predominant genetic predisposition to ASD seems beyond doubt, its exact nature remains unclear. In the context of social cognition disorders and the basis of ASD, the oxytocinergic and vasopresynergic systems arouse great interest among researchers. The aim of the present study was to analyze gene expression levels for oxytocin and vasopressin receptors, as well as CD38 protein and oxytocinase, in the context of the clinical picture of autism spectrum disorders. The study included 90 people, of whom 63 were diagnosed with ASD based on anamnesis, mental status testing, and the ADOS-2 protocol. The results obtained in the presented study indicate that the balance between the levels of expression of the CD38 gene and the oxytocinase gene plays a key role in the risk and clinical presentation of ASD. In a hypothetical scenario, an imbalance in the expression of CD38 and LNPEP could potentially lead to alterations in the concentrations of oxytocin and vasopressin. At the same time, the most frequently studied genes-AVPR1a and OXTR-seem to be at best of marginal importance for the risk of ASD.

Comparison of cultures and 16S rRNA sequencing for identification of bacteria in two-stage revision arthroplasties: preliminary report.

BACKGROUND: The use of a prefabricated spacer in two-stage revision arthroplasty remains one of the few surgery strategies for infected-joint arthroplasty treatment, despite the many unidentified microorganisms in the infected joint replacements reported in some recent studies. The aim of this prospective survey was to investigate if the sonication followed by polymerase chain reaction (PCR) can improve bacterial identification on the surfaces of prefabricated spacers and if the systemic laboratory mediators of infection and positive microbiological results can take a role of predictive factors of infection and clinical failures in 2-years follow-up. METHODS: Thirteen patients with prosthetic joint infection were investigated. Bacterial culture and deoxyribonucleic acid (DNA) sequencing were used to detect bacteria on the surface of prefabricated spacers removed during the second stage of revision arthroplasty. The results of pre- and intraoperative culture and DNA sequencing were compared. Minimum follow-up was 2 years. RESULTS: The result of tissue cultures in second-stage revision arthroplasties revealed positive results in 15 % of patients with Coagulase-negative Staphylococci (CNS) growth. Bacterial DNA was found in over 90 % of patients with negative synovial fluid culture. Positive PCR results revealed potential pathogenic bacteria and species of human and environmental microflora with low virulence. Clinical failures at final follow-up were recorded in 2 (16.6 %) patients. CONCLUSION: The lack of clinical signs of infection, negative culture of preoperative joint aspirate, and intraoperative specimens do not exclude the presence of bacteria on the surfaces of spacers. The positive results of sonication and molecular tests should be interpreted as real pathogenicity factors in the light of the clinical and laboratory data, especially for patients with immunodeficiency. We confirmed our previous results that sonication followed by PCR and sequencing improved bacterial identification.

Analysis of the TID-I and TID-L Splice Variants' Expression Profile under In Vitro Differentiation of Human Mesenchymal Bone Marrow Cells into Osteoblasts.

Bone formation is a complex process regulated by a variety of pathways that are not yet fully understood. One of the proteins involved in multiple osteogenic pathways is TID (DNAJA3). The aim of this work was to study the association of TID with osteogenesis. Therefore, the expression profiles of the TID splice variants (TID-L, TID-I) and their protein products were analyzed during the proliferation and differentiation of bone marrow mesenchymal stromal cells (B-MSCs) into osteoblasts. As the reference, the hFOB1.19 cell line was used. The phenotype of B-MSCs was confirmed by the presence of CD73, CD90, and CD105 surface antigens on ~97% of cells. The osteoblast phenotype was confirmed by increased alkaline phosphatase activity, calcium deposition, and expression of ALPL and SPP1. The effect of silencing the TID gene on the expression of ALPL and SPP1 was also investigated. The TID proteins and the expression of TID splice variants were detected. After differentiation, the expression of TID-L and TID-I increased 5-fold and 3.7-fold, respectively, while their silencing resulted in increased expression of SPP1. Three days after transfection, the expression of SPP1 increased 7.6-fold and 5.6-fold in B-MSCs and differentiating cells, respectively. Our preliminary study demonstrated that the expression of TID-L and TID-I changes under differentiation of B-MSCs into osteoblasts and may influence the expression of SPP1. However, for better understanding the functional association of these results with the relevant osteogenic pathways, further studies are needed.

Identification of silent prosthetic joint infection: preliminary report of a prospective controlled study.

PURPOSE: We will test the hypothesis that ultrasound supported by polymerase chain reaction (PCR) could improve bacterial identification in non-infected prosthetic joint loosening. The aim was to detect bacterial species in non-infected prosthetic joint loosening using ultrasound and 16S rRNA gene sequencing. METHODS: A total of 16 patients (11 women and five men) aged 46-80 years (mean age 65.7) with diagnosed knee or hip implant loosening (mean implant survival of 102.1 months) were investigated. Bacterial culture and DNA sequencing were used to detect bacteria on the surface of failed implants removed during revision arthroplasty. The results of pre- and intraoperative culture and DNA sequencing were compared. Histopathological analysis was also performed. RESULTS: The number of positive cultures rises with a higher level of C-reactive protein (CRP). The results of the cultures from synovial fluid obtained through joint aspiration were consistent with sonicates from components of prostheses in 12 cases (75%). Bacterial DNA was found in 90% of patients with negative synovial fluid culture. PCR revealed two or more bacterial species, often of the same genus: Ralstonia pickettii, Pseudomonas spp., Brevibacterium spp., Lactobacillus spp., Propionibacterium spp. and Staphylococcus spp.These are micro-organisms present in the environment or on the human body and often associated with compromised immunity. CONCLUSIONS: The ultrasound procedure followed by PCR and sequencing improve bacterial identification in silent prosthetic joint infection. The lack of clinical signs of infection and negative preoperative and intraoperative cultures do not exclude the presence of micro-organisms on the implants.

Collagen and elastin differences in vulvar tissue of women with lichen planus, lichen sclerosus and healthy women.

OBJECTIVES: Lichen sclerosus and lichen planus are two debilitating dermatoses. Their etiology remains unknown. Skin changes resulting from these disorders are important to understand, so we can provide targeted treatment to patients. We examined the differences in collagen (COL1A1, COL1A2, COL3A1, COL5A1, COL5A2, COL5A3) and elastin (ELN) expression between vulvar tissue of women with lichen planus, lichen sclerosus and healthy women. MATERIAL AND METHODS: Vulvar tissue was taken from areas affected by lichen planus or lichen sclerosus. In healthy controls, we biopsied vulva at five and eight o'clock in a standardized manner. The tissue was simultaneously sent for pathological and genetic analysis. When either lichen planus or sclerosus or healthy tissue was confirmed by pathologist, we processed the genetic sample. RNA was isolated, transcribed and gene expression was analyzed using Real Time Custom Panel 96-16 and LightCycler 480 Probe Master. Kolmogorov-Smirnov test was employed to determine if the data on the population show normal distribution. For genes with normal distribution, t-Test was employed and for those lacking normality, we used Mann-Whitney 1-tail test. The threshold for p-value was set less than 0.05. RESULTS: Thirty-nine vulvar samples were examined. The mean expression of COL1A1 was 11.13, COL1A2 was 6.72, COL3A1 was 8.43, COL5A1 was 11.91, COL5A2 was 10.62 and COL5A3 was 12.79. The mean expression of elastin (ELN) was 13,13. We found statistically significant difference in expression of collagen (COL1A2) and elastin (ELN) between healthy controls and patients with lichen planus (p = 0.4). We did not find differences for other genes (p < 0.05). CONCLUSIONS: Collagen and elastin are differentially expressed between patients with lichen planus and healthy controls.

Polymorphisms in Oxytocin and Vasopressin Receptor Genes as a Factor Shaping the Clinical Picture and the Risk of ASD in Males.

Autism spectrum disorders (ASD) are a heterogeneous group of disorders affecting virtually every population, regardless of their ethnic or socioeconomic background. Their pathogenesis is multifactorial, based on interactions between genetic and environmental factors. The key symptom of ASD are deficits in social communication, which are the basis of many difficulties in everyday functioning. The aim of the presented study was to analyze the clinical picture of social cognition deficits in boys with autism spectrum disorders and to relate its elements with the frequency of alleles of selected polymorphisms within the oxytocin receptor (OXTR) and vasopressin receptor 1A (AVPR1A) genes. The study included 58 boys with IQ > 90, who were divided into two groups based on a confirmed or excluded ASD diagnosis based on the DSM-5 and ICD-10 criteria and then using the ADOS-2 protocol. The results indicated that polymorphism rs10877969 (T) within the AVPR1a gene was the only one to show a statistically significant association with a higher risk of autism spectrum disorders and has an impact on clinical presentation in the ADOS-2 study, primarily in terms of the social affect subscale. Polymorphisms in the OXTR gene showed no significant association with ASD risk and severity of autistic traits in the ADOS-2 study. In the group of people with ASD and those who are neurotypical, the rs53572 (A) genotype in the OXTR gene significantly increased the severity of the clinical picture of social cognition disorders in reading mind in the eyes test (RMiE) and empathy quotient (EQ) studies.

Expression of heat shock proteins HSPA1A, HSPA1B and TP53 in vulval lichen planus and vulval lichen sclerosus.

OBJECTIVES: Heat shock proteins (HSPs) are proteins involved in protein folding and maturation. HSP expression is induced by heat shock or other stressors including cellular damage and hypoxia. The major groups, which are classified based on their molecular weight, include HSP27, HSP40, HSP60, HSP70, HSP90, and large HSP (HSP110 and glucose-regulated protein 170). The comparison of heat shock proteins and TP53 expression is yet not well studied in both vulval lichen sclerosus and lichen planus. Our aim was to assess the HSP and TP53 gene expression in women suffering from LS or LP and compare it within these groups and also healthy controls. MATERIAL AND METHODS: The inclusion criteria were willingness to donate vulval biopsies, not currently or in the prior two weeks received any local nor systemic treatment for vulval disorder, age > 18 years old. The exclusion criteria were lack of consent, current vaginal infection confirmed with microbiological studies, current local or systemic treatment for vulval disease. 45 consecutive women were recruited into the study. All appropriate vulval samples were process by genetic analysis. RESULTS: The mean expression (± SD) of HPSA1A for controls was 5.52 ± 3.18, for LS was 7.44 ± 2.16 and for LP was 7.89 ± 2.48. The mean expression (± SD) of HPSA1B for controls was 6.54 ± 3.41, for LS was 9.94 ± 6.88 and for LP was 9.43 ± 2.31. The mean expression (± SD) of TP53 for controls was 9.11 ± 1.14, for LS was 9.94 ± 1.27 and for LP was 10.41 ± 2.00. HSPA1A expression was 3,8 higher in women with lichen sclerosus than in control group. CONCLUSIONS: Heat shock protein-70 is more often expressed in LS than in healthy controls. HSP-70 not only supports tumor growth and metastasis, but on the other hand mat help to develop immune-driven treatment strategies.

The role of selected polymorphisms in regulation of gene CD38 expression and their effect on the clinical picture of autism spectrum disorders - preliminary study. / Rola wybranych polimorfizmów w regulacji ekspresji genu CD38 i ich wplyw na ryzyko oraz obraz kliniczny zaburzen ze spektrum autyzmu.

OBJECTIVES: Clinical effects observed in cases of oxytocin deficiency can also manifest themselves in disorders of mechanisms responsible, for example, for its secretion. For oxytocin, this function is played by - among others - the cluster of differentiation antigen 38 (CD38). Existing literature along with the correlation between protein CD38 and oxytocin secretion raise interest in the context of their possible relation to the clinical picture and development of the autism spectrum disorders (ASD). The aim of the study was to analyze the correlations between polymorphisms rs3796863 and rs6449197 in gene CD38, the level of gene expression and the clinical picture and the risk of ASD diagnosis. METHODS: The study included 59 individuals with the mean age of 15.05 years with IQ > 90. The participants were divided into two groups: the studied group consisting of 37 persons with confirmed ASD diagnoses and the control group including 22 neurotypical individuals. Diagnosis verification was carried out via the ADOS-2 protocol. RESULTS: The comparative analysis with the standardized population based on the 1000Genomes database with the presence of clinically significant intensification of ASD traits showed the correlation of alleles "T" of polymorphisms rs3796863 and rs6449197, which are more frequent in the general population and are treated as "wild". In the inter-group analysis, this type of dependency was weaker, and the genotype of the control group was somehow intermediate between the studied group and the standardized population. In the ΔΔCt analysis, the normalized value of the relative expression level of gene CD38 showed that in the studied group the expression level was around 1.1-1.2 times higher than in the control group. CONCLUSIONS: The obtained results show that a significant correlation with the severity of autism spectrum disorder traits is mainly observed in the carriers of wild variants of the studied polymorphisms, in which the related increase in the expression level of gene CD38 is also observed.

Analysis of miRNAs in Osteogenesis imperfecta Caused by Mutations in COL1A1 and COL1A2: Insights into Molecular Mechanisms and Potential Therapeutic Targets.

Osteogenesis imperfecta (OI) is a group of connective tissue disorders leading to abnormal bone formation, mainly due to mutations in genes encoding collagen type I (Col I). Osteogenesis is regulated by a number of molecules, including microRNAs (miRNAs), indicating their potential as targets for OI therapy. The goal of this study was to identify and analyze the expression profiles of miRNAs involved in bone extracellular matrix (ECM) regulation in patients diagnosed with OI type I caused by mutations in COL1A1 or COL1A2. Primary skin fibroblast cultures were used for DNA purification and sequence analysis, followed by analysis of miRNA expression. Sequencing analysis revealed mutations of the COL1A1 or COL1A2 genes in all OI patients, including four previously unreported. Amongst the 40 miRNAs analyzed, 9 were identified exclusively in OI cells and 26 in both OI patients and the controls. In the latter case, the expression of six miRNAs (hsa-miR-10b-5p, hsa-miR-19a-3p, hsa-miR-19b-3p, has-miR-204-5p, has-miR-216a-5p, and hsa-miR-449a) increased, while four (hsa-miR-129-5p, hsa-miR-199b-5p, hsa-miR-664a-5p, and hsa-miR-30a-5p) decreased significantly in OI cells in comparison to their expression in the control cells. The identified mutations and miRNA expression profiles shed light on the intricate processes governing bone formation and ECM regulation, paving the way for further research and potential therapeutic advancements in OI and other genetic diseases related to bone abnormality management.

mRNA Expression of thrombospondin 1, 2 and 3 from proximal to distal in human abdominal aortic aneurysm - preliminary report.

Abdominal aortic aneurysm is a process involving the disruption and reconstruction of the extracellular matrix and the apoptosis of smooth muscle cells under the strong influence of the immune system. Thrombospondins are proteins that influence a wide range of cell-matrix interactions. While THBS1 and THBS2 are widely studied, the effects of THBS3 on extracellular matrix and vascular cells are poorly understood. Additionally, it is not known whether expression of these genes' changes along the aneurysm tissue. Here we analyzed the expression of THBSs mRNA isolated from the harvested tissues along the aneurysm divided into three zones based on their morphology. Total mRNA was isolated from 13 male patients undergoing scheduled open aortic repair, with each aneurysm divided into a proximal part, an aneurysm bag, and a distal part with border tissue as a control. Two step real-time PCR analysis with random hexamers was performed, which allowed the detection of significantly increased expression of all analyzed thrombospondins, especially THBS3, at the control tissue. Overexpression of THBSs may have a destabilizing effect on the structure of the extracellular matrix by affecting both the matrix producing cells and by inhibiting the activity of matrix proteins.

Searching for new molecular markers for cells obtained from abdominal aortic aneurysm.

The aim of the study was to investigate specific potential markers for cells obtained from three layers of human AAA divided into three segments along the AAA based on morphological differences. The isolated cells were compared to control commercial cell types from healthy human abdominal aortas. For each type of aortic layer, three specimens from 6 patients were compared. Total RNA was isolated from 36 cell cultures for gene expression profiling and potential new cytometry markers were typed. Isolated cells were analyzed by flow cytometry by using fluorochrome-conjugated antibodies to markers: CNN1, MYH10, ENG, ICAM2, and TEK. The relative expression of 45 genes in primary cell cultures and control lines was analyzed. Statistically significant differences were found in the expression of most of the analyzed genes between individual layers and control lines. Based on relative expression, antibodies were selected for flow cytometry. Gene expression profiles allowed to select new potential cytometry markers: CNN1, MYH10, MYOCD, ENG, ICAM2, TEK. However, none of the tested markers seems to be optimal and characteristic for a specific layer of AAA.

Expression gradient of metalloproteinases and their inhibitors from proximal to distal segments of abdominal aortic aneurysm.

Abdominal aortic aneurysm refers to abnormal, asymmetric distension of the infrarenal aortic wall due to pathological remodelling of the extracellular matrix. The distribution of enzymes remodelling the extracellular matrix and their expression patterns in the affected tissue are largely unknown. The goal of this work was to investigate the expression profiles of 20 selected genes coding for metalloproteinases and their inhibitors in the proximal to the distal direction of the abdominal aortic aneurysm. RNA samples were purified from four lengthwise fragments of aneurysm and border tissue obtained from 29 patients. The quantities of selected mRNAs were determined by real-time PCR to reveal the expression patterns. The genes of interest encode collagenases (MMP1, MMP8, MMP13), gelatinases (MMP2, MMP9), stromelysins (MMP3, MMP7, MMP10, MMP11, MMP12), membrane-type MMPs (MMP14, MMP15, MMP16), tissue inhibitors of metalloproteinases (TIMP1, TIMP2, TIMP3, TIMP4), and ADAMTS proteinases (ADAMTS1, ADAMTS8, and ADAMTS13). It was found that MMP, TIMP, and ADAMTS are expressed in all parts of the aneurysm with different patterns. A developed aneurysm has such a disturbed expression of the main participants in extracellular matrix remodelling that it is difficult to infer the causes of the disorder development. MMP12 secreted by macrophages at the onset of inflammation may initiate extracellular matrix remodelling, which, if not controlled, initiates a feedback loop leading to aneurysm formation.

Presence of human papilloma virus in Caucasian women living in the central Europe diagnosed with vulvar intraepithelial neoplasia.

OBJECTIVES: The role of human papilloma virus (HPV) in the development of cancerous states of female reproductive tract has been widely debated. However, the information about presence of HPV in the Caucasian women living in the central Europe diagnosed with vulvar intraepithelial neoplasia (VIN) is missing. So far, no recommendation was made to complete HPV detection in time of vulvar biopsy or after the results of positive VIN are obtained. We aimed to assess the presence of HPV in women with vulvar intraepithelial neoplasia diagnosed at the Department of Gynecology, Obstetrics and Oncological Gynecology in Bytom, Poland. MATERIAL AND METHODS: The retrospective examination of 120 consecutive vulvar biopsies obtained from women with persistent vulvar itching was done. Only patients with diagnosis of VIN were included in the further analysis. HPV DNA was detected using HPV Linear Array Genotyping Test including 14 HPV genotypes (16, 18, 31, 33, 35, 39, 45, 51, 52, 56, 58, 59, 66, 68). RESULTS: Out of 120 vulvar samples retrieved, 18 women were positive for VIN, including15 usual VIN (uVIN) and three differentiated type (dVIN ). 10 samples were eligible for DNA detection. HPV DNA was found in two women with uVIN (HPV 16 and 51). CONCLUSIONS: It is advisable to recommend HPV genotyping in women with VIN, regardless of their age and histologic type. The incidence of HPV infection in Caucasian women from the central Europe with VIN should be further studied.

Serologic investigations in children with inflammatory bowel disease and food allergy.

The aim of the study was the evaluation of frequency and titre of IgA ASCA and IgG ASCA and p-ANCA, c-ANCA in children with IBD and occurrence of ASCA antibodies in relation to coexistence of FA. Patients and methods. The study comprised 95 children at the ages of 2 to 18 years. The diagnosis of IBD was established on the basis of Porto criteria. Tests of blood serum were performed in all children: IgA and IgG ASCA, p-ANCA, c-ANCA using ELISA method. Results. IgE-dependent FA was found in 32.5% children with UC and in 21% with CD. We did not observe any relation between the occurrence of FA and the frequency and ASCA titre. p-ANCA were significantly more frequent in the group of children with UC. The occurrence of ASCA antibodies was observed in 73.7% of children with CD, 17.5% with UC and almost 30% with allergic colitis. Conclusions. Patients with CD and the presence of ASCA revealed a significantly more frequent localization of lesions within the small bowel and a tendency towards older age. We observed a connection between the occurrence of antibodies and the examined mutations of gene NOD2/CARD15.

Blocking angiogenesis with peptides that inhibit the activity of procollagen C-endopeptidase.

Procollagen C-endopeptidase (BMP-1) is one of two key enzymes crucial for conversion of fibrillar procollagens to self-assembling collagen monomers. Recently, we have reported inhibition of the largest variant of BMP-1, a recombinant mammalian tolloid (mTld) in vitro, on procollagen type I using peptides with amino acid sequences in chordin conserved across different species. Here, we tested the same peptides as potent blockers of angiogenesis ex vivo in cultured rings of rat aorta, in vivo in chick embryos, and in vitro in cell cultures. Our results revealed that the peptides inhibited the angiogenic activity in rat aorta explants at micromolar concentrations; they also blocked blood vessel growth in chick embryos. The peptides were also tested on three types of human cells, e.g., umbilical vein endothelium, skin fibroblasts, and tumor HT-1080 cells. Since the three types of cells proliferated at a significantly lower rate or did not proliferate at all, we conclude that the anti-angiogenic effect observed in rat aorta ring explants and in chick embryos was related to inhibition of cell proliferation. In conclusion, we showed the ability to inhibit angiogenesis by blocking the activity of procollagen C-endopeptidase. The results strongly indicate crucial role(s) of this metalloproteinase in the formation of new blood vessels and maintenance of their growth.

[Changes of mRNAs encoding alternatively spliced variants of procollagen C-endopeptidase in leiomyomas uteri depending on the phase of menstrual cycle and in postmenopausal women]. / Zmiany mRNA wariantów alternatywnego skladania C-endopeptydazy prokolagenu w miesniakach macicy w zaleznosci od fazy cyklu miesiecznego i u kobiet w okresie klimakterium.

UNLABELLED: Leiomyoma uteri is a monoclonal tumour of the uterus muscle layer. It is characterized by excessive, abnormal growth of extracellular matrix. The collagen types I and III are the major components of extracellular matrix. Removal of the C-propeptides in procollagens type I, II, and III by procollagen C-endopeptidase leads to spontaneous self-assembly of collagen fibrils. Thus, the procollagen C-endopeptidase is a key regulator of extracellular matrix production, its quality, and other developmental processes including angiogenesis. OBJECTIVE: The objective of this study was to analyze the three alternatively spliced variants of the BMP1 gene product including the procollagen C-endopeptidase, in leiomyoma uteri tumors in comparison to normal myometrium in women being in first or second menstrual phase or in time of climacterium. MATERIAL AND METHODS: In the study we analyzed samples from the control and cancerous tissues from 52 women. Expression of the three alternatively spliced variants of BMP1 gene transcript were assayed by RT-PCR, following densytometric analysis. Statistical significance (p < or = 0,05) of the observed differences was assessed with the use of Anova test, Least Significance Different Test, and Student's T-test. RESULTS: Analysis of RT-PCR products revealed the presence of all alternatively spliced variants of BMP-1 mRNA and the changes in their alternative splicing intensity, depending on the phase of menstrual cycle or postmenopausal state. In the second phase of the cycle, in control tissue, the expression level of BMP-1 variant decreased when compared to women in I phase of the cycle or postmenopausal women. In the tumour, postmenopausal women showed increased expression of BMP-1at, when compared with women in the second phase of the cycle. The presence of mTLD in the tumour tissue at II phase of the cycle and in postmenopausal state was less strong when compared to women at I phase of the cycle. In control tissue this type of change was not observed. The BMP-1/HIS was present at higher level in control tissue, during II phase of the menstrual cycle and in postmenopausal state, whereas in the tumour tissue its lowest level was at II phase of the cycle. CONCLUSIONS: Regulation of alternative splicing of mRNA for procollagen C-endopeptidase in leiomyomas and myometrium depends mainly on the hormonal status of women.

Roles of microRNAs in abdominal aortic aneurysm pathogenesis and the possibility of their use as biomarkers.

Abdominal aortic aneurysm (AAA) is a dangerous condition involving pathological dilation of the aortic wall. Due to the asymptomatic course of this disease and the dangerous consequences of its rupture, it is important to identify its specific bio-markers expressed as early as possible. Different expression profiles of microRNAs (miRNAs) were detected in patients diagnosed with AAA. MicroRNAs are small non-coding RNA molecules that regulate the expression of other genes at the translation stage. miRNAs affecting translation can lead to abnormal remodeling of extracellular matrix, inhibition of the cell cycle, cell aging or intensified inflammation. This review summarizes current knowledge about the role of microRNAs in the context of formation and development of abdominal aorta aneurysm and the possibility of using some miRNAs as bio-markers, and also provides basic information about miRNAs and aneurysms.

Specific inhibition of procollagen C-endopeptidase activity by synthetic peptide with conservative sequence found in chordin.

Procollagen C-endopeptidase (BMP-1) and N-endopeptidase (ADAMTS-2) are key enzymes for correct and efficient conversion of fibrillar procollagens to their self assembling monomers. Thus, they have an essential role in building and controlling the quality of extracellular matrices (ECMs). Here, we tested inhibition of activity of the largest variant of BMP-1, a recombinant mammalian tolloid (mTld), in vitro by three synthetic peptides with conservative amino-acid sequences found in chordin using procollagen type I as a substrate. We also verified the specific action of best inhibitory 16 amino-acid peptide in the procollagen type I cleavage assay with the use of ADAMTS-2 (procollagen N-endopeptidase). Subsequently, we determined the critical residues and minimal sequence of six amino acids in the original 16 amino-acid peptide required to maintain the inhibitory potential. Studies on the interactions of 6 and 16 amino acid long peptides with the enzyme revealed their binding to non-catalytic, regulatory domains of mTld; the inhibitory activity was not due to the competition of peptides with the substrate for the enzyme active center, because mTld did not cleave the peptides. However, in the presence of mTld both peptides underwent cyclization by disulfide bond formation. Concluding, we have shown that procollagen C-endopeptidase may be specifically blocked via its non-catalytic domains by synthetic peptide consisting of 6 amino acids in the sequence found in highly conservative region of chordin. Thus, we hypothesize that the 6 amino-acid peptide could be a good candidate for anti-fibrotic drug development.

Two novel COL1A1 mutations in patients with osteogenesis imperfecta (OI) affect the stability of the collagen type I triple-helix.

Osteogenesis imperfecta (OI) is a bone dysplasia caused by mutations in the COL1A1 and COL1A2 genes. Although the condition has been intensely studied for over 25 years and recently over 800 novel mutations have been published, the relation between the location of mutations and clinical manifestation is poorly understood. Here we report missense mutations in COL1A1 of several OI patients. Two novel mutations were found in the D1 period. One caused a substitution of glycine 200 by valine at the N-terminus of D1 in OI type I/IV, lowering collagen stability by 50% at 34 degrees C. The other one was a substitution of valine 349 by phenylalanine at the C-terminus of D1 in OI type I, lowering collagen stability at 37.5 degrees C. Two other mutations, reported before, changed amino residues in D4. One was a lethal substitution changing glycine 866 to serine in genetically identical twins with OI type II. That mutated amino acid was near the border of D3 and D4. The second mutation changed glycine 1040 to serine located at the border of D4 and D0.4, in a proband manifesting OI type III, and lowered collagen stability at 39 degrees C (2 degrees C lower than normal). Our results confirm the hypothesis on a critical role of the D1 and D4 regions in stabilization of the collagen triple-helix. The defect in D1 seemed to produce a milder clinical type of OI, whereas the defect in the C-terminal end of collagen type caused the more severe or lethal types of OI.

[Molecular characteristics of leiomyoma uteri based on selected compounds of the extracellular matrix]. / Molekularna charakterystyka guzów leiomyoma uteri na przykladzie wybranych skladników macierzy pozakomórkowej.

Leiomyoma is a monoclonal benign tumor. It is often located in the muscle layer of the uterus in women of reproductive age. Its growth is accelerated by pregnancy and hormonal therapy. Its growth also depends on the concentration of sex hormones. Growth factors and cytokines may also participate in the formation of leiomyomas. The modulation of mitotic activity and abnormal extracellular matrix production are key elements of tumor growth. Elements of the TGFbeta superfamily are crucial factors in the proliferation of neoplasmic cells. TGF-beta1 and -beta3 stimulate the synthesis of various components of the extracellular matrix, but they also down-regulate the synthesis of proteinases which degrade the matrix, often leading to excessive overdeposition of connective tissue. Collagen types 1 and 3 are the main structural components of the extracellular matrix. The biosynthesis of collagens requires, among others, the action of procollagen C-endopeptidase, a protein of the BMP-1/mTLD subfamily. BMP-1/mTLD-like proteinases remove the carboxyl propeptides of procollagens 1, 2, and 3. Removal of the C-propeptides decreases the solubility of procollagens about 1000-fold to a concentration critical for their spontaneous self-assembly to collagen fibrils. Different substrates of BMP-1/mTLD are prolysyl oxidase, gamma2 chain of prolaminin, procollagen type VII, miostatin, dentin matrix protein 1, and perlekan. Due to the activation of various substrates by BMP-1/mTLDs, they are important regulators of the production of the extracellular matrix and its quality as well as of antiangiogenic responses by producing a factor from the basal membrane compound called perlekan. The BMP-1/mTLDs influence the formation of dorsal ventral patterning in embryos by releasing BMP-2/4 from the inhibitory protein chordin. Another aspect is induction of the development of muscle and neural tissue by activation of GDF8 and GDF11 as well as the regulation of growth and cell proliferation by releasing TGF-beta1 and -beta3 from latent complexes. Another yet poorly understood aspect is the evolution of neoplastic cells based on other than molecular genetic mechanisms. The detectable karyotype anomalies in tumor cells constitute just 40%. Therefore in this review the possible roles of extracellular matrix compounds and regulatory factors in the pathology of leiomyoma are discussed.