Hydroxychloroquine Enhances Efferocytosis and Modulates the Gut Microbiome in Mice With Pristane-induced Lupus

Background An important contributor to the pathogenesis of the autoimmune disease systemic lupus erythematosus (SLE) is the impaired clearance of apoptotic cells (efferocytosis) and increasing evidence implicates microbiota dysbiosis as another important player. The disease-modifying antirheumatic drug hydroxychloroquine (HCQ) is frequently prescribed for the treatment of SLE. Here, we evaluate changes in efferocytosis and the gut microbiome in mice with pristane-induced lupus (PIL) before and after HCQ administration.Methods PIL mice were studied with or without HCQ and/or resveratrol (RESV). Efferocytosis was determined in RAW 264.7 cells and peritoneal macrophages from mouse ascites fluid. The gut microbiome was analyzed using Illumina sequencing targeting the 16S ribosomal DNA gene (rDNA) amplicon sequencing.Results Both HCQ and RESV enhanced efferocytosis. HCQ also significantly suppressed ascites production of proinflammatory cytokines interleukin (IL)-6 and tumor necrosis factor alpha (TNF-α). The Firmicutes/Bacteriodetes (F/B) ratio was significantly decreased in PIL mice compared with untreated controls (p<0.05). The F/B ratio in PIL mice was increased by HCQ alone and significantly increased by HCQ combined with RESV. Alpha- and beta-diversity differed between mice administered RESV and those that were not. Viable counts of lactic acid bacteria Lactobacillaceae, Lactobacillus and Lactobacillales were increased by RESV plus HCQ treatment.Conclusions In this study, HCQ and RESV enhanced efferocytosis in both RAW 264.7 cells and peritoneal macrophages of PIL mice and suppressed IL-6 and TNF-α production in ascites fluid. The pristane-induced reduction in the F/B ratio was restored by HCQ treatment. SLE treatment should consider the role of the gut microbiome.

An Alarming Mimicry of Intra-Abdominal Infections: Acute Appendiceal Diverticulitis

A 65-year-old woman presented with three days of colicky abdominal pain. Abdominal imaging illustrated small bowel enteritis, ascites in both paracolic gutters, and incidental hepatic steatosis. Although ascites fluid demonstrated high neutrophil count consistent with peritonitis and the patient received adequate antibiotics, she clinically deteriorated. Subsequent exploratory laparotomy revealed necrotic appendix and multiple intra-abdominal abscesses. Histopathology showed acute suppurative appendicitis with multiple other intact small diverticula, indicating likely perforation of inflamed appendiceal diverticula with subsequent abscess formation and abdominal peritonitis. This case highlights the importance of ascites fluid analysis and continued clinical correlation, especially in cases of rare entities with atypical presentations.

Induction of Ascites in Mice

Smaller animals such as rats, mice, hamsters, and guinea pigs are usually poor choices for polyclonal antibody production because only small volumes of serum can be obtained. This problem can be reduced by inducing the formation of ascites in mice, which can provide up to 10 mL of ascites fluid from a single animal. Antibody titers in ascites fluids are almost as high as serum titers.

Comparison of Diagnostic Accuracy of Portal Serum Ascitic Albumin Gradient (SAAG) with Ascitic Fluid Total Protien (AFTP) Differentiating Portal Hypertension from Non-Portal in Patients with Ascites

Introduction: Although majority of the cases of ascites have cirrhosis, there are 15% patients where there is a non-hepatic cause of fluid retention like malignancy, congestive heart failure and tuberculous peritonitis. Ascites is the most common complication of cirrhosis that leads to hospital admission. Objective:To compare the diagnostic Accuracy of Serum Ascitic Albumin Gradient (SAAG) and Ascitic Fluid Total Proteins in patients with ascites by taking Ultrasound abdomen & Pelvis as gold standard. There are international studies on the accuracy of SAAG in determining cause of ascites but not much local data. Additionally, SAAG is not widely used in our setup. The results of this study will add to the existing knowledge and will help in the diagnosis and better management of these patients. Material & Methods: A cross sectional validation study was conducted in the department of General Medicine, DHQTH, Dera Ismail Khan from 29th April to 29th Oct, 2019. Diagnostic Ascitic fluid was aspirated from the peritoneal cavity and ascitic fluid was sent to hospital laboratory for total protein and albumin. Blood was taken at the same time and was send to the hospital laboratory for the serum albumin. SAAG was calculated by subtracting ascitic albumin value from the serum albumin value. Both, Ascitic fluid total protein and SAAG values was documented in the proforma. Ultrasound Abdomen & Pelvis was done on each patient with special instruction for radiologist to comment upon Portal Vein diameter and any changes in its diameter with respiration. Results: As per comparison Of SAAG with ultrasound in detecting ascites, sensitivity was 36.26%, specificity was 75%, PPV was 84.62%, NPV was 23.68% and accuracy was 44.35%. P Value was 0.299. As per comparison of AFTP with ultrasound in detecting ascites, sensitivity was 33.33%, specificity was 59.34%, PPV was 17.78%, NPV was 77.14% and accuracy was 53.91%. P value was 0.513. Conclusion:SAAG exhibits that patients with ascites fluid possess the basis of portal hypertension. Thus we have come to this conclusion that SAAG can effectively enhance the diagnostic value of ascites fluid tests and therefore its classification can be considered to be a novel standard in the analysis of ascites fluid. Keywords: Diagnostic Accuracy, Ascites Volume, Ascitic Albumin Gradient (SAAG), Ascitic Fluid Total Proteins (AFTP)

Penetration of Isavuconazole in Ascites Fluid of Critically Ill Patients

Fungal peritonitis is a life-threatening condition which is not only difficult to diagnose, but also to treat. Following recent guidelines, echinocandins and azoles are the recommended antimycotics for the management of intra-abdominal Candida spp. infections, with a favor for echinocandins in critically ill patients. However, the new extended spectrum triazole isavuconazole also has a broad spectrum against Candida spp. Data on its target-site penetration are sparse. Therefore, we assessed isavuconazole concentrations and penetration ratios in ascites fluid of critically ill patients. Obtaining of Isavuconazole plasma and ascites fluid levels as well penetration ratios using paracentesis in critically ill patients. Isavuconazole concentrations were quantified in human plasma and ascites by a liquid chromatography/tandem mass spectrometry (LC-MS/MS) method. Isavuconazole concentrations in plasma and ascites fluid were measured in sixteen critically ill patients. Isavuconazol levels in ascites fluid (1.06 µg/mL) were lower than plasma levels (3.08 µg/mL). Penetration ratio was 36%. In two out of sixteen patients, Candida spp., in detail C. glabrata and C. tropicalis, could be isolated. Cmax/MIC Ratio in plasma of 560 for C. glabrata and 2166 for C. tropicalis could be observed. Following our results, isavuconazole penetrates into ascites. Successful treatment in Candida spp. peritonitis depends on pathogen susceptibility.

Clinical and Bacteriological features of spontaneous bacterial peritonitis in Malagasy patients with Cirrhosis

Background: Spontaneous bacterial peritonitis (SBP) represent frequent and serious complications in cirrhosis patients with ascites. Our aim was to describe the clinical and bacteriological characteristics of SBP in Madagascar.Methods: This is a 21-month prospective study between January 2018 and October 2019, including hospitalized patients with cirrhosis, with clinical and biological symptoms of SBP.Results: Thirty-three patients were included. The mean age was 48.09 ± 13.55 years (extremes: 19 – 78 years), the sex ratio was 3.12. Abdominal pain (55%), fever (36%), diarrhea (6%), hepatic encephalopathy (18%) are the most common symptoms. Gastrointestinal bleeding (18.18%) was the main risk factor to SBP. SBP was community-acquired in 87.88% of cases. A culture of ascites fluid was positive for 9 patients (27.27%). The infectious agents found were Escherichia coli (12.10%), Klebsiella pneumoniae (3%), Pseudomonas (3%), Streptococcus mitis (9.1%). Escherichia coli were wild with one case resistant to Ceftriaxone. The Klebsiella were multidrug resistant. The other two pathogens did not show resistance. After antibiotic therapy adapted to the antibiogram, healing was observed in 26 patients (78.78%). Seven patients (21.22%) died from various complications. All deceased patients had bacteria identified in ascites fluid.Conclusion: SBP defined according to clinical and biological criteria is apparently sterile in the majority of cases. Gram-negative bacteria were the major pathogens involved in SBP in cirrhotic patients. Escherichia coli and streptococcus were the most common pathogen isolated. Bacteriological study is essential to adapt antimicrobial to multidrug-resistant bacteria.

Hypocalcemia in sepsis: analysis of the subcellular distribution of Ca2+ in septic rats and LPS/TNF-α-treated HUVECs

Introduction: Hypocalcemia has been widely recognized in sepsis patients. However, the cause of hypocalcemia in sepsis is still not clear, and little is known about the subcellular distribution of Ca2+ in tissues during sepsis. Methodology: We measured the dynamic change in Ca2+ levels in body fluid and subcellular compartments, including the cytosol, endoplasmic reticulum and mitochondria, in major organs of cecal ligation and puncture (CLP)-operated rats, as well as the subcellular Ca2+ flux in HUVECs which treated by endotoxin and cytokines. Results: In the model of CLP-induced sepsis, the blood and urinary Ca2+ concentrations decreased rapidly, while the Ca2+ concentration in ascites fluid increased. The Ca2+ concentrations in the cytosol, ER, and mitochondria were elevated nearly synchronously in major organs in our sepsis model. Moreover, the calcium overload in CLP-operated rats treated with calcium supplementation was more severe than that in the non-calcium-supplemented rats but was alleviated by treatment with the calcium channel blocker verapamil. Similar subcellular Ca2+ flux was found in vitro in HUVECs and was triggered by lipopolysaccharide (LPS)/TNF-α. Conclusions: Ca2+ influx from the blood into the intercellular space and Ca2+ release into ascites fluid may cause hypocalcemia in sepsis and that this process may be due to the synergistic effect of endotoxin and cytokines.