The Role of Thrombin in Proliferative Vitreoretinopathy

PURPOSE. To determine the role of thrombin in the development of proliferative vitreoretinopathy (PVR). METHODS. Vitreous was collected from patients undergoing a vitrectomy (macular holes and puckers, n ¼ 11 [controls]; retinal detachment without PVR development following vitrectomy, n ¼ 15 [RRD1]; retinal detachment with PVR development within 6 months after vitrectomy, n ¼ 11 [RRD2]; and established PVR, n ¼ 14 [PVR]). Thrombin activity in vitreous was determined using a thrombin-specific chromogenic substrate. ARPE-19 cells were stimulated with 83 diluted vitreous samples in the presence and absence of hirudin. The samples were analyzed at t ¼ 0 and t ¼ 24 hours for the presence of 27 cytokines/ chemokines and growth factors using a multiplex approach. In comparable studies, ARPE-19 cells were stimulated for 2 hours, and mRNA expression levels for CCL2, CXCL8, GMCSF, IL6, and PDGFB were determined by real-time quantitative (RQ)-PCR. RESULTS. Thrombin activity was significantly (P < 0.05) higher in vitreous of the PVR group compared to the other groups. Proliferative vitreoretinopathy vitreous stimulated the production of chemokine (C-C motif) ligand (CCL)2, chemokine (C-X-C motif) ligand (CXCL)8, granulocyte-macrophage colony-stimulating factor (GM-CSF), IL-6, and plateletderived growth factor (PDGF)-BB by ARPE-19 to significantly (P < 0.05) higher levels than vitreous from the RRD1 and RRD2 groups. These effects of PVR vitreous were significantly (P < 0.05) reduced by hirudin. These data were confirmed by mRNA studies. CONCLUSIONS. Thrombin activity is increased in vitreous of patients with established PVR and is involved in the activation of proinflammatory and profibrotic pathways in RPE cells. Inhibition of thrombin activity may therefore represent a potential treatment option for proliferative vitreoretinopathy

Immunological profiling in long COVID:overall low grade inflammation and T-lymphocyte senescence and increased monocyte activation correlating with increasing fatigue severity

Background: Many patients with SARS-CoV-2 infection develop long COVID with fatigue as one of the most disabling symptoms. We performed clinical and immune profiling of fatigued and non-fatigued long COVID patients and age- and sex-matched healthy controls (HCs). Methods:Long COVID symptoms were assessed using patient-reported outcome measures, including the fatigue assessment scale (FAS, scores ≥22 denote fatigue), and followed up to one year after hospital discharge. We assessed inflammation-related genes in circulating monocytes, serum levels of inflammation-regulating cytokines, and leukocyte and lymphocyte subsets, including major monocyte subsets and senescent T-lymphocytes, at 3-6 months post-discharge. Results: We included 37 fatigued and 36 non-fatigued long COVID patients and 42 HCs. Fatigued long COVID patients represented a more severe clinical profile than non-fatigued patients, with many concurrent symptoms (median 9 [IQR 5.0-10.0] vs 3 [1.0-5.0] symptoms, p&lt;0.001), and signs of cognitive failure (41%) and depression (&gt;24%). Immune abnormalities that were found in the entire group of long COVID patients were low grade inflammation (increased inflammatory gene expression in monocytes, increased serum pro-inflammatory cytokines) and signs of T-lymphocyte senescence (increased exhausted CD8+ TEMRA-lymphocytes). Immune profiles did not significantly differ between fatigued and non-fatigued long COVID groups. However, the severity of fatigue (total FAS score) significantly correlated with increases of intermediate and non-classical monocytes, upregulated gene levels of CCL2, CCL7, and SERPINB2 in monocytes, increases in serum Galectin-9, and higher CD8+ T-lymphocyte counts. Conclusion: Long COVID with fatigue is associated with many concurrent and persistent symptoms lasting up to one year after hospitalization. Increased fatigue severity associated with stronger signs of monocyte activation in long COVID patients and potentially point in the direction of monocyte-endothelial interaction. These abnormalities were present against a background of immune abnormalities common to the entire group of long COVID patients.</p

Association of monocyte HLA-DR expression over time with secondary infection in critically ill children: a prospective observational study

An impaired immune response could play a role in the acquisition of secondary infections in critically ill children. Human leukocyte antigen-DR expression on monocytes (mHLA-DR) has been proposed as marker to detect immunosuppression, but its potential to predict secondary infections in critically ill children is unclear. We aimed to assess the association between mHLA-DR expression at several timepoints and the change of mHLA-DR expression over time with the acquisition of secondary infections in critically ill children. In this prospective observational study, children < 18 years with fever and/or suspected infection (community-acquired or hospital-acquired) were included at a paediatric intensive care unit in the Netherlands. mHLA-DR expression was determined by flow cytometry on day 1, day 2–3 and day 4–7. The association between delta-mHLA-DR expression (difference between last and first measurement) and secondary infection was assessed by multivariable regression analysis, adjusted for age and Paediatric Logistic Organ Dysfunction-2 score. We included 104 patients at the PICU (median age 1.2 years [IQR 0.3–4.2]), of whom 28 patients (27%) developed a secondary infection. Compared to 93 healthy controls, mHLA-DR expression of critically ill children was significantly lower at all timepoints. mHLA-DR expression did not differ at any of the time points between patients with and without secondary infection. In addition, delta-mHLA-DR expression was not associated with secondary infection (aOR 1.00 [95% CI 0.96–1.04]). Conclusions: Our results confirm that infectious critically ill children have significantly lower mHLA-DR expression than controls. mHLA-DR expression was not associated with the acquisition of secondary infections.What is Known:• An impaired immune response, estimated by mHLA-DR expression, could play an essential role in the acquisition of secondary infections in critically ill children.• In critically ill children, large studies on the association of mHLA-DR expression with secondary infections are scarce.What is New:• Our study confirms that critically ill children have lower mHLA-DR expression than healthy controls.• mHLA-DR expression and change in mHLA-DR was not associated with the acquisition of secondary infection

Serum Levels of Leptin As Marker For Patients At High Risk of Gastric Cancer

Background: Serological screening for gastric cancer (GC) may reduce mortality. However, optimal serum markers for advanced gastric precursor lesions are lacking.Aim: To evaluate in a case-control study whether serum leptin levels correlate with intestinal metaplasia (IM) and can serve as a tool to identify patients at high risk for GC.Materials and Methods: Cases were patients with a previous diagnosis of IM or dysplasia, controls were patients Without such a diagnosis. All patients underwent endoscopy. Fasting serum was collected for the measurement of leptin, pepsinogens I/II, gastrin, and Helicobacter pylori. Receiver operating characteristic (ROC) Curves and their area under the curve (AUC) were provided to compare serum leptin levels with other serological markers.Results: One hundred nineteen cases and 98 controls were included. In cases, the median leptin levels were 116.6 pg/mL versus 81.9 pg/mL, in controls (p = .01). After adjustment for age, sex and BMI, leptin levels remained higher in cases than in controls (p &lt;.005). In multivariate analysis, male sex (p = .002), age (Conclusions: High leptin levels are associated with an increased risk of IM. Moreover, serum leptin levels are a significant independent marker for the presence of IM. However, in combination with the serological test for pepsinogen I the additional value of serum leptin levels is rather limited.</p

Encapsulating peritoneal sclerosis is associated with T-cell activation

Background Encapsulating peritoneal sclerosis (EPS) is an excessive fibrotic response of the peritoneum that may occur after long-term peritoneal dialysis (PD). The underlying pathophysiology is poorly understood, but involvement of peritoneal inflammatory T helper 1 cells may be pivotal. Methods Soluble interleukin-2 receptor alpha (sCD25) concentration was measured as a marker for T-cell activation in serum and ascites from EPS patients and various control patient groups. Peritoneal biopsies were stained for the presence of T cells, and T cells isolated from ascites of EPS patients were characterized in detail for differentiation status and cytokine expression. Results Serum sCD25 concentrations are significantly and specifically increased in EPS patients compared with haemodialysis, PD and predialysis patients. Peritoneal effluent of stable PD patients contains very low levels of sCD25, while sCD25 levels in ascites of EPS patients are high and indicative of local production. In the years preceding the diagnosis of EPS, the serum sCD25 concentrations increased while remaining at stable levels in control PD patients. The peritoneum and ascites of EPS patients showed a significant influx of T cells with relatively increased numbers of CD4+ T cells. These T cells were fully differentiated and displayed a T helper 1 cell type with a pro-inflammatory cytokine profile. Conclusions Increased serum sCD25 concentrations and peritoneal lymphocytosis in EPS patients indicate the involvement of activated T cells in the pathophysiology of excessive fibrosis

Human decidual tissue contains differentiated CD8+ effector-memory T cells with unique properties

During pregnancy, maternal lymphocytes at the fetal-maternal interface play a key role in the immune acceptance of the allogeneic fetus. Recently, CD4(+)CD25(bright) regulatory T cells have been shown to be concentrated in decidual tissue, where they are able to suppress fetus-specific and nonspecific immune responses. Decidual CD8(+) T cells are the main candidates to recognize and respond to fetal HLA-C at the fetal-maternal interface, but data on the characteristics of these cells are limited. In this study we examined the decidual and peripheral CD8(+) T cell pool for CD45RA, CCR7, CD28, and CD27 expression, using nine-color flow cytometry. Our data demonstrate that decidual CD8(+) T cells mainly consist of differentiated CD45RA(-)CCR7(-) effector-memory (EM) cells, whereas unprimed CD45RA(+)CCR7(+) naive cells are almost absent. Compared with peripheral blood EM CD8(+) T cells, the decidual EM CD8(+) T cells display a significantly reduced expression of perforin and granzyme B, which was confirmed by immunohistochemistry of decidual tissue sections. Interestingly, quantitative PCR analysis demonstrates an increased perforin and granzyme B mRNA content in decidual EM CD8(+) T cells in comparison with peripheral blood EM CD8(+) T cells. The presence of high levels of perforin and granzyme B mRNA in decidual EM T cells suggests that decidual CD8(+) T cells pursue alternative means of EM cell differentiation that may include a blockade of perforin and granzyme B mRNA translation into functional perforin and granzyme B proteins. Regulation of decidual CD8(+) T cell differentiation may play a crucial role in maternal immune tolerance to the allogeneic fetu

Encapsulating peritoneal sclerosis is associated with T-cell activation

Background Encapsulating peritoneal sclerosis (EPS) is an excessive fibrotic response of the peritoneum that may occur after long-term peritoneal dialysis (PD). The underlying pathophysiology is poorly understood, but involvement of peritoneal inflammatory T helper 1 cells may be pivotal. Methods Soluble interleukin-2 receptor alpha (sCD25) concentration was measured as a marker for T-cell activation in serum and ascites from EPS patients and various control patient groups. Peritoneal biopsies were stained for the presence of T cells, and T cells isolated from ascites of EPS patients were characterized in detail for differentiation status and cytokine expression. Results Serum sCD25 concentrations are significantly and specifically increased in EPS patients compared with haemodialysis, PD and predialysis patients. Peritoneal effluent of stable PD patients contains very low levels of sCD25, while sCD25 levels in ascites of EPS patients are high and indicative of local production. In the years preceding the diagnosis of EPS, the serum sCD25 concentrations increased while remaining at stable levels in control PD patients. The peritoneum and ascites of EPS patients showed a significant influx of T cells with relatively increased numbers of CD4+ T cells. These T cells were fully differentiated and displayed a T helper 1 cell type with a pro-inflammatory cytokine profile. Conclusions Increased serum sCD25 concentrations and peritoneal lymphocytosis in EPS patients indicate the involvement of activated T cells in the pathophysiology of excessive fibrosis

Presentation_1_Immunological profiling in long COVID: overall low grade inflammation and T-lymphocyte senescence and increased monocyte activation correlating with increasing fatigue severity.pdf

BackgroundMany patients with SARS-CoV-2 infection develop long COVID with fatigue as one of the most disabling symptoms. We performed clinical and immune profiling of fatigued and non-fatigued long COVID patients and age- and sex-matched healthy controls (HCs).MethodsLong COVID symptoms were assessed using patient-reported outcome measures, including the fatigue assessment scale (FAS, scores ≥22 denote fatigue), and followed up to one year after hospital discharge. We assessed inflammation-related genes in circulating monocytes, serum levels of inflammation-regulating cytokines, and leukocyte and lymphocyte subsets, including major monocyte subsets and senescent T-lymphocytes, at 3-6 months post-discharge.ResultsWe included 37 fatigued and 36 non-fatigued long COVID patients and 42 HCs. Fatigued long COVID patients represented a more severe clinical profile than non-fatigued patients, with many concurrent symptoms (median 9 [IQR 5.0-10.0] vs 3 [1.0-5.0] symptoms, p24%). Immune abnormalities that were found in the entire group of long COVID patients were low grade inflammation (increased inflammatory gene expression in monocytes, increased serum pro-inflammatory cytokines) and signs of T-lymphocyte senescence (increased exhausted CD8+ TEMRA-lymphocytes). Immune profiles did not significantly differ between fatigued and non-fatigued long COVID groups. However, the severity of fatigue (total FAS score) significantly correlated with increases of intermediate and non-classical monocytes, upregulated gene levels of CCL2, CCL7, and SERPINB2 in monocytes, increases in serum Galectin-9, and higher CD8+ T-lymphocyte counts.ConclusionLong COVID with fatigue is associated with many concurrent and persistent symptoms lasting up to one year after hospitalization. Increased fatigue severity associated with stronger signs of monocyte activation in long COVID patients and potentially point in the direction of monocyte-endothelial interaction. These abnormalities were present against a background of immune abnormalities common to the entire group of long COVID patients.</p