Cytokine profile in women with threatened miscarriage and intestinal dysbiosis

Hypothesis/Aims of study: Systemic endotoxinemia resulting from intestinal dysbiosis activates the Th1 immune response and excessive production of proinflammatory cytokines, which can cause abortion. This study was aimed at assessing interleukin levels in women with the threat of miscarriage and exploring their dependence on intestinal microbiocenosis and the level of endotoxinemia. Study design, materials and methods: The study involved 87 women aged 18 to 43 years in pregnancy from six to 22 weeks. The main group consisted of 50 women with a threatened miscarriage. The control group included 37 women with normal pregnancy. A qualitative and quantitative analysis of intestinal microbiocenosis was performed by real-time PCR, with endotoxin and interleukin levels evaluated using conventional methods. Results: In patients of the main group, intestinal dysbiosis of grade I was detected in 30 % (n = 15), of grade II in 46 % (n = 23), and of grade III in 24 % (n = 12) of cases. In the control group, intestinal microflora disorders corresponded to grade I dysbiosis in 67.6 % (n = 25) and grade II dysbiosis in 32.4 % (n = 12) of cases, there being no cases of severe dysbiosis revealed. The level of endotoxin in pregnant women of the main group was 0.57 0.02 nmol / ml and was classified as increased. The endotoxin level in the blood serum of pregnant women of the control group was 0.34 0.02 nmol / ml and was characterized as low (p 0.001). A strong correlation was found (r = 0.8, p 0.001) between the grade of intestinal dysbiosis and the level of endotoxinemia. The cytokine profile in patients of the main group was characterized by increased concentrations of pro-inflammatory cytokines (IL-1 4.9 1.6 pg / ml, IL-6 4.8 1.5 pg / ml) and a decreased concentration of anti-inflammatory cytokine (IL-10 18.0 4.5 pg / ml), when compared to the control group (IL-1 1.8 0.2 pg / ml, IL-6 2.1 0.2 pg / ml, IL-10 30.3 4.4 pg/ml). There were a statistically significant moderate positive correlation between the level of endotoxinemia and the concentration of pro-inflammatory cytokines and a weak negative correlation between the endotoxin level and the concentration of anti-inflammatory IL-10. Conclusion: Endotoxinemia, which occurs as a result of intestinal microflora dysbiosis and activates pro-inflammatory pathways, can really be as a triggering factor in the pathogenesis of miscarriage in pregnant women.

Impact of Endotoxins in Gelatine Hydrogels on Chondrogenic Differentiation and Inflammatory Cytokine Secretion In Vitro

Gelatine methacryloyl (GelMA) hydrogels are widely used in studies aimed at cartilage regeneration. However, the endotoxin content of commercially available GelMAs and gelatines used in these studies is often overlooked, even though endotoxins may influence several cellular functions. Moreover, regulations for clinical use of biomaterials dictate a stringent endotoxin limit. We determined the endotoxin level of five different GelMAs and evaluated the effect on the chondrogenic differentiation of equine mesenchymal stromal cells (MSCs). Cartilage-like matrix production was evaluated by biochemical assays and immunohistochemistry. Furthermore, equine peripheral blood mononuclear cells (PBMCs) were cultured on the hydrogels for 24 h, followed by the assessment of tumour necrosis factor (TNF)-α and C–C motif chemokine ligand (CCL)2 as inflammatory markers. The GelMAs were found to have widely varying endotoxin content (two with >1000 EU/mL and three with <10 EU/mL), however, this was not a critical factor determining in vitro cartilage-like matrix production of embedded MSCs. PBMCs did produce significantly higher TNF-α and CCL2 in response to the GelMA with the highest endotoxin level compared to the other GelMAs. Although limited effects on chondrogenic differentiation were found in this study, caution with the use of commercial hydrogels is warranted in the translation from in vitro to in vivo studies because of regulatory constraints and potential inflammatory effects of the content of these hydrogels.

Endotoxemia in metabolic syndrome and some mechanisms of its development

The aim. Assessment of the relationship between the occurrence of endotoxemia and impaired resorption function of the small intestine, arrhythmia of the motility of the stomach and intestines, and excessive bacterial growth in the metabolic syndrome (MS).Materials and methods. 62 patients with MS were examined. The average age was 48.62+3.75 years. The motor function of the gastrointestinal tract was studied using peripheral electrogastroenterocolography. To assess the absorption processes in the small intestine, stress tests with glucose and d-xylose were used. Blood endotoxin level, quantitative and& qualitative composition of parietal microbiota of small intestine were determined using an Agilent gas chromatograph with mass-selective and flame-ionization detectors (Agilent Technologies, USA).Results. 82.9% of the examined patients with MS showed clinical signs of intestinal damage. The electrical activity of& the& small intestine departments in the postprandial period was low in 70% of patients. The electrical activity of& the& colon on an empty stomach was not changed, and after a food load was reduced. Discoordination of motility is observed between small intestine and the colon, aggravated after food stimulation. A significant decrease in the rhythm of contractions is observed at the frequencies of the jejunum, ileum, and colon both on an empty stomach and in the postprandial period, which indicates a weakening of propulsive bowel contractions in patients with MS. In patients with MS, an increase in& the& absorption of glucose and d-xylose was revealed in comparison with the group of healthy individuals. In patients with MS, excessive bacterial growth in small intestine is observed mainly due to conditionally pathogenic microbiota strains. According to the results of the study of blood endotoxin level in patients with MS, a significant increase was revealed in comparison with the control group. Correlation analysis showed a moderate negative relationship between the level of endotoxin and electrical activity of small intestine, between the level of endotoxin and the ratio of the ratio of& the& colon to the ileum. A moderate positive relationship was established between the degree of endotoxemia increase and glucose absorption in small intestine.Conclusion. A study of the level of endotoxin and indicators of excessive bacterial growth, motor evacuation and resorption functions of small intestine revealed important pathogenetic patterns regarding the contributing role of the latter in& the development of endotoxemia in patients with MS.

Impact of endotoxins in gelatine hydrogels on chondrogenic differentiation and inflammatory cytokine secretion in vitro

Gelatine methacryloyl (GelMA) hydrogels are widely used in studies aiming at cartilage regeneration. However, the endotoxin content of commercially available GelMAs and gelatines used in these studies is often overlooked, even though endotoxins may influence several cellular functions. Moreover, regulations for clinical use of biomaterials dictate a stringent endotoxin limit.We determined the endotoxin level of five different GelMAs and evaluated the effect on the chondrogenic differentiation of equine mesenchymal stromal cells (MSCs). Cartilage-like matrix production was evaluated by biochemical assays and immunohistochemistry. Furthermore, equine peripheral blood mononuclear cells (PBMCs) were cultured on the hydrogels for 24 hours, followed by the assessment of tumour necrosis factor (TNF)-α and C-C motif chemokine ligand (CCL)2 as inflammatory markers.The GelMAs were found to have widely varying endotoxin content (two with >1000 EU/ml and three with <10 EU/ml), however, this was not a critical factor determining in vitro cartilage-like matrix production of embedded MSCs. PBMCs did produce significantly higher TNF-α and CCL2 in response to the GelMA with the highest endotoxin level compared to the other GelMAs.Although limited effects on chondrogenic differentiation were found in this study, caution with the use of commercial hydrogels is warranted in the translation from in vitro to in vivo studies because of regulatory constraints and potential inflammatory effects of the content of these hydrogels.