Role of increased neutrophil extracellular trap formation on acute kidney injury in COVID-19 patients.

Background: A strong association between elevated neutrophil extracellular trap (NET) levels and poor clinical outcomes in patients with coronavirus infection 2019 (COVID-19) has been reported. However, while acute kidney injury (AKI) is a common complication of COVID-19, the role of NETs in COVID-19-associated AKI is unclear. We investigated the association between elevated NETs and AKI and the prognostic role of NETs in COVID-19 patients. Methods: Two representative markers of NETs, circulating nucleosomes and myeloperoxidase-DNA, were measured in 115 hospitalized patients. Serum levels of interleukin [IL]-6, monocyte chemotactic protein-1 [MCP-1], plasma von Willebrand factor (vWF) and urinary biomarkers of renal tubular damage (ß2-microglobulin [ß2M] and kidney injury molecule 1 [KIM-1]) were measured. Results: AKI was found in 43 patients (37.4%), and pre-existing chronic kidney disease (CKD) was a strong risk factor for AKI. Higher circulating NET levels were a significant predictor of increased risk of initial ICU admission, in-hospital mortality (adjusted HR 3.21, 95% CI 1.08-9.19) and AKI (OR 3.67, 95% CI 1.30-10.41), independent of age, diabetes, pre-existing CKD and IL-6 levels. There were strong correlations between circulating nucleosome levels and urinary KIM-1/creatinine (r=0.368, p=0.001) and ß2M (r=0.218, p=0.049) levels. NETs were also strongly closely associated with serum vWF (r = 0.356, p<0.001), but not with IL-6 or MCP-1 levels. Conclusions: Elevated NETs were closely associated with AKI, which was a strong predictor of mortality. The close association between NETs and vWF may suggest a role for NETs in COVID-19-associated vasculopathy leading to AKI.

Increased viability of endometrial cells by in vitro treatment with di-(2-ethylhexyl) phthalate.

Based on the findings of several reports that have shown an increased plasma level of di-(2-ethylhexyl) phthalate (DEHP) in women with endometriosis, the present study was designed to evaluate whether in vitro treatment with DEHP can increase viability of endometrial cells. Utilizing 3-(4,5-dimethyl-2-thiazolyl)-2,5-diphenyltetrazolium bromide assay, fluorescent activated cell sorter analysis, and microscopic evaluation after Hoechst staining, we revealed that in vitro treatment with DEHP leads to increased viability of Ishikawa cells as well as endometrial stromal cells in serum-free condition and following exposure to hydrogen peroxide, which suggests that exposure to phthalate might play a role in the establishment of endometriosis.

Increased expression of p21-activated kinase in adenomyosis.

The present study was designed to evaluate whether the expression of p21-activated kinase 1 (Pak1), which recently has been shown to be increased in the eutopic endometrium of women with endometriosis, is also increased in adenomyotic nodules as well as in the eutopic endometrium in women with adenomyosis. Comparative immunohistochemical study using a monoclonal antihuman Pak1 antibody revealed that the expression of Pak1 is increased in the eutopic endometrium of women with adenomyosis during the secretory phase, along with its increased expression in adenomyotic nodules, suggesting a possible role of Pak1 in the establishment of adenomyosis.

Increased expression of glutathione by estradiol, tumor necrosis factor-alpha, and interleukin 1-beta in endometrial stromal cells.

PROBLEM: The intracellular antioxidant system, based on glutathione (GSH), plays a key role in endometrial detoxification reactions and has been proposed to be involved in the pathogenesis endometriosis. This study was designed to evaluate whether estradiol (E(2)) and proinflammatory cytokines have any effects on expression of glutathione in endometrial stromal cells (ESCs). METHOD OF STUDY: Glutathione levels were measured utilizing high-performance liquid chromatography following in vitro culture and treatment of ESCs with estradiol, tumor necrosis factor-alpha (TNF-alpha) and interleukin 1-beta (IL-1beta). RESULTS: The GSH level in E(2) (10(-8) m) treatment group was significantly higher than in the control group at 48 h (P < 0.05). In vitro treatment of ESCs with TNF-alpha 10 ng/mL as well as E(2) (10(-8) m) plus TNF-alpha 10 ng/mL for 48 hr also led to a significant increase in GSH level (P < 0.05; P < 0.05, respectively). Both IL-1beta 10 ng/mL and E(2) (10(-8) m) plus IL-1beta 10 ng/mL for 48 hr increased GSH level significantly (P < 0.05; P < 0.05, respectively) as well. CONCLUSIONS: These findings might suggest that increased production of estradiol and proinflammatory cytokines in the peritoneal cavity possibly leads to the establishment of endometriosis through increased level of GSH.

Down-regulation of p21-activated kinase 1 by progestin and its increased expression in the eutopic endometrium of women with endometriosis.

BACKGROUND: P21-activated kinase 1 (Pak1) integrates various signaling pathways that are vital to cell survival and function. This study was performed to evaluate whether sex steroids may regulate the expression of Pak1 in endometrial cells as well as whether its expression is increased in the eutopic endometrium of women with endometriosis. METHODS: Following in vitro estradiol (E(2)) and/or medroxyprogesterone acetate (MPA) treatment of Ishikawa cells and endometrial stromal cells (ESCs), Pak1 protein was analyzed utilizing western blot analysis and immunocytochemistry. Immunohistochemistry was performed to evaluate Pak1 immunoreactivity semiquantitatively in women with endometriosis and in controls. To assess the role of Pak1 on endometrial cell viability, crystal violet assay was performed following transfection of Ishikawa cells with Pak1 small interfering RNA (siRNA). RESULTS: In vitro treatment with E(2) plus MPA or MPA alone led to a significant decrease of Pak1 protein in Ishikawa cells and ESCs (both P < 0.05 versus control). Immunohistochemistry also revealed that Pak1 protein is significantly decreased during the secretory phase in both epithelial and stromal cells in the control subjects (P < 0.001 and P < 0.01, respectively). The immunoreactivity of Pak1 in glandular cells was significantly increased in the eutopic endometrium of women with endometriosis compared with the controls during the secretory phase (P < 0.01). Crystal violet assay has shown that transfection of Ishikawa cells with Pak1 siRNA led to a significant decrease of cellular viability (P < 0.05). CONCLUSIONS: These findings suggest that Pak1 is down-regulated by progesterone during the secretory phase in normal endometrium and increased Pak1 activity during the secretory phase might lead to establishment of endometriosis.