Peritoneal Dialysis Fluid Research Articles

Performance evaluation of UF-4000 body fluid mode for detection of bacteria in body fluids

BackgroundMicrobiological analysis of body fluids (BF) provides important information for diagnosis of infection. We evaluated the analytical performance of bacterial count by UF-4000 BF mode for ascitic, cerebrospinal, pleural, synovial and continuous ambulatory peritoneal dialysis fluids compared to classical microbiological procedure (direct Gram staining and culture). Materials and methodsFor the 1,734 BF analyzed, distribution of UF-4000 bacterial count was analyzed according to the level of growth culture and results were compared using Mann-Whitney test. ROC curves analysis allowed to define the best cut-off value to predict or exclude positive culture for each type of BF. ResultsUF-4000 bacterial counts were significantly lower in sterile than in infected BFs (p < 0.00001) and correlated with the level of growth on culture. The ROC curves of bacteria/μL and culture positivity yielded area under the curve >0.80 for each type of BF. Optimal cut-offs were chosen with excellent statistical parameters (sensitivity ranging from 0.70 to 0.86, specificity from 0.78 to 0.98, negative predictive value >0.95 and Youden index >0.55). ConclusionFor BF, UF-4000 bacterial count correlate with culture results and is a discriminative method enhancing detection of microbiological etiology. It could be used as a screening method based on the cut-off values proposed in this study.

Is icodextrin use feasible in a peritoneal dialysis patient undergoing 18 F-FDG PET/CT scan?

18 F-FDG PET/CT scan is a useful diagnostic tool in patients with neoplasia or inflammatory diseases for further evaluation. Due to interference of glucose with the cellular uptake of the 18 F-FDG tracer via glucose transporter, withhold of any glucose source several hours before imaging is mandatory. This is also the case in peritoneal dialysis patients where glucose-containing peritoneal dialysis fluid cannot be used prior to 18 F-FDG PET/CT scan. Whether the same hold true for icodextrin is not known. We describe two patients with metastatic carcinomas while on peritoneal dialysis on an icodextrin-containing regimen, in whom icodextrin was not discontinued before 18 F-FDG PET/CT scan and where accurate diagnosis of metastatic lesions as well as in one case a simultaneous tunnel infection could be made. Our observation suggests that there is no significant interference of icodextrin with the metabolism of 18 F-FDG and so it is feasible to continue an established icodextrin-containing regimen in peritoneal dialysis patients in case of 18 F-FDG PET/CT scan.

Endoglin aggravates peritoneal fibrosis by regulating the activation of TGF-β/ALK/Smads signaling.

Background: Peritoneal fibrosis (PF) is an intractable complication in patients on long-term peritoneal dialysis (PD). Transforming growth factor-β (TGF-β) is a key pro-fibrogenic factor involved in PD-associated PF, and endoglin, as a coreceptor for TGF-β, plays a role in balancing the TGF-β signaling pathway. Here, we investigated whether endoglin could be a potential therapeutic target for PF. Methods: In vivo, we established PF model in SD rats by daily intraperitoneal injection of peritoneal dialysis fluids (PDF) containing 4.25% glucose for 6 weeks and downregulated endoglin expression by tail vein injection of AAV9-ENG on day 14 to assess the effect of endoglin on peritoneal morphology and markers related to fibrosis, angiogenesis, and epithelial-mesenchymal transition (EMT). In vitro, we treated human peritoneal mesothelial cells (HPMCs) transfected with ENG siRNA in high glucose medium to explore the potential mechanism of endoglin in PF. Results: Compared to control group, continuous exposure to biologically incompatible PDF induced exacerbated PF, accompanied by a significant increase in endoglin expression. Conversely, knockdown of endoglin ameliorated peritoneal injury characterized by increased peritoneal thickening and collagen deposition, angiogenesis, as well as EMT. Consistently, HPMCs cultured in high glucose medium underwent the EMT process and exhibited over-expression of fibronectin, collagen type I, vascular endothelial growth factor (VEGF), whereas these aforementioned alterations were alleviated after ENG siRNA transfection. In addition, we also found that ENG siRNA inhibited TGF-β-induced phosphorylation of Smad2/3 and Smad1/5/9 in HPMCs treated with high glucose (HG). Conclusion: Our findings confirmed for the first time that endoglin exacerbated PF by regulating the activation of TGF-β/ALK/Smads signaling, which will provide a novel potential therapeutic target in PF.

Spontaneous peritoneal dialysis catheter expulsion: A case series.

There are a significant number of mechanical complications related the peritoneal dialysis (PD) catheter and early diagnosis is key. Between them, the spontaneous extrusion of the catheter is quite uncommon but it should be included among the possible complications of the technique, given that it can be related to frequent situations in PD patients like abdominal wall issues, past immunosuppression or PD fluid leakage. In 25 years our Unit had three cases, probably related to peritoneal fluid leakage, exit site infection and past steroid treatment. Careful examination of abdominal wall preimplantation, adequate break-in period to allow maturation and early detection of exit site infection to start treatment in a timely fashion could help to prevent this rare complication. The complete extrusion of the catheter does not preclude from continuing PD treatment.

Assessment of solution stability and drug release properties of liposomal curcumin in peritoneal dialysis fluid and its synergistic antibacterial activity with vancomycin

Infectious Peritonitis associated with recurrent and persistent infections is the major cause of technique failure of life-sustaining peritoneal dialysis (PD) therapy used for treating patients with end-stage renal failure (ESRF). The infections -if diagnosed timely- are usually resolved with empiric antibiotic treatment. However, the treatment of infections associated with drug resistant bacteria or fungal infections remains challenging, poses a very serious problem to the health of PD patients, and is the leading cause of mortality and morbidity. Therefore, the development of alternate approaches other than antibiotics are required to keep up with the constantly changing and increasing multiple drug resistance (MDR) of bacterial strains. Curcumin exhibits anti-oxidative/anti-inflammatory potential and remarkable wound healing properties. It also has broad-spectrum anti-bacterial/anti-biofilm effects and its synergistic antibacterial activity with several antibiotics (including those used clinically for the treatment of PD associated infections) is well proven against variety of pathogenic infections including MDR strains. However, poor water-solubility of curcumin (<0.125 mg/L) remains a major barrier to achieve desirable bioavailability and further limit its therapeutic efficacy. Among the various drug carrier strategies, liposomal encapsulation has been found to be most promising mode to remarkably enhance the therapeutic index of curcumin by facilitating its gastric absorption, protecting its enzymatic degradation and allowing its slow release over time for maximum benefits. However, to date, the potential of liposomal curcumin has not been explored for the treatment of PD associated infections. Starting our efforts in this direction, a comprehensive assessment of solution stability and drug release properties of curcumin-loaded liposomal vesicles has been demonstrated in peritoneal dialysis fluid. The results clearly indicated an excellent solution stability of curcumin loaded inside liposome and the formulation affirms the regulated curcumin release enhancing the bioavailability. Further, we evaluated the synergistic antibacterial efficacy of curcumin-loaded liposomal formulation with vancomycin against the Staphylococcus aureus and Escherichia coli – pathogens (known for causing intra-peritoneal infections during PD). This synergy strategy resulted in outstanding improvement in bacterial inhibition efficacy of liposomal curcumin at significantly lowered doses. The results of the present study propose lipocurcumin as a potent antibiotic additive to vancomycin to cure PD-infections caused by antibiotic-resistant bacteria.

P180 Is gene sequencing only way forward?

Poster session 2, September 22, 2022, 12:30 PM - 1:30 PM IntroductionFungal peritonitis (FP) associated with peritoneal dialysis (PD) is an uncommon complication. Candida peritonitis accounts for 70%-90%. A total of 55 cases of Aspergillus peritonitis have been reported from 1968 to 2019 with A. fumigatus and A. niger as the most common isolates.FP is associated with high mortality and often results in discontinuation of PD and switching to hemodialysis (HD).ObjectivesTo identify and report the pathogen from peritoneal abscess.Methods and ResultsA 54-year-old post-menopausal lady, a known diabetic, hypertensive with chronic kidney disease (CKD) on PD for 2 years presents with acute onset breathlessness. She was dialyzed and improved. During her hospital stay, she developed a fever which was managed with intravenous antibiotics, piperacillin-tazobactam, teicoplanin, and fluconazole. The PD was bacterial culture negative. Patient improved on HD and was discharged.A total of 10 days later she was readmitted due to complaints of fever. Her PD fluid analysis was clear with high leucocyte count. The same was sent for the presence of acid-fast bacilli and aerobic bacterial culture, both of which were negative. Her blood was sent for automatic aerobic bacterial culture which yielded no growth. Patient developed pain in abdomen and PD catheter was removed. Abdominal Computed Tomography (CT) imaging revealed a thick-walled intra-peritoneal collection in the left subdiaphragmatic region. In view of persistent fever, antibiotics were changed to meropenem, metronidazole, and amikacin. Pus aspirated was received for aerobic bacterial culture and fungal culture. KOH—Calcofluor examination revealed septate fungal hyphae. Bacterial culture yielded Enterococcus species sensitive to penicillin and gentamicin. Patient was treated with amphotericin B and HD. Repeat aspirate also revealed septate fungal hyphae. However, on both occasions fungal cultures were negative. She improved symptomatically and was discharged.Sample was sent to Post Graduate Institute of Medical Education and Research, Chandigarh for the identification of the fungal pathogen directly from the sample using Gene sequencing. The pathogen was identified as A. niger.Conclusion Aspergillus peritonitis in a PD patient though rare is associated with high mortality and morbidity. They are often associated with concurrent or prior bacterial peritonitis. Treatment with antibiotics often predisposes to fungal peritonitis. Clinical presentation is similar to bacterial peritonitis, a high index of suspicion is warranted to establish the diagnosis. Identification of fungal pathogen to rule out species inherently resistant to certain classes of antifungals is critical for successful management. This is possible only by molecular methods, especially in culture-negative cases. In this case of culture-negative invasive fungal infection, the preliminary diagnosis was based on microscopic evidence of two consecutive samples. Gene sequencing helped us to confirm the diagnosis.

Effect of Plasma Resuscitation with Adjunctive Peritoneal Resuscitation on Hepatic Blood Flow and End-Organ Damage after Hemorrhagic Shock.

Intestinal injury from resuscitated hemorrhagic shock (HS) disrupts intestinal microvascular flow and causes enterocyte apoptosis, intestinal barrier breakdown, and injury to multiple organs. Fresh frozen plasma (FFP) resuscitation or directed peritoneal (DPR) resuscitation protect endothelial glycocalyx, improve intestinal blood flow, and alleviate intestinal injury. We postulated that FFP plus DPR might improve effective hepatic blood flow (EHBF) and prevent associated organ injury (liver, heart). Anesthetized Sprague-Dawley rats underwent HS (40% mean arterial pressure, 60 minutes) and were randomly assigned to groups (n = 8 per group): Sham; crystalloid resuscitation (CR; shed blood + 2 volumes CR); DPR (intraperitoneal 2.5% peritoneal dialysis fluid); FFP (shed blood + 1 vol IV FFP); FFP + DPR. EHBF was measured at postresuscitation timepoints. Organ injury was evaluated by serum ELISA (fatty acid-binding protein [FABP]-1 [liver], FABP-3 [heart], Troponin-I [heart], and Troponin-C [heart]) and hematoxylin and eosin. Differences were evaluated by 1-way ANOVA and 2-way repeated-measures ANOVA. CR resuscitation alone did not sustain EHBF. FFP resuscitation restored EHBF after resuscitation (2 hours, 3 hours, and 4 hours). DPR resuscitation restored EHBF throughout the postresuscitation period but failed to restore serum FABP-1 VS other groups. Combination FFP + DPR rapidly and sustainably restored EHBF and decreased organ injury. CR and DPR alone had elevated organ injury (FABP-1 [hepatocyte], FABP-3 [cardiac], and Troponin-I/C), whereas FFP or FFP + DPR demonstrated reduced injury at 4 hours after resuscitation. HS decreased EHBF, hepatocyte injury, and cardiac injury as evidenced by serology. FFP resuscitation improved EHBF and decreased organ damage. Although DPR resuscitation resulted in sustained EHBF, this alone failed to decrease hepatocyte or cardiac injury. Combination therapy with DPR and FFP may be a novel method to improve intestinal and hepatic blood flow and decrease organ injury after HS/resuscitation.

Inactivation of the Complement Lectin Pathway by Candida tropicalis Secreted Aspartyl Protease-1

Candida tropicalisis an opportunistic fungal pathogen and is one of the most frequently isolated non-albicans species. It can cause localised as well as invasive systemic infections particularly in immunocompromised patients. Increased resistance to common anti-fungal drugs is an emerging problem. In order to establish disseminated infections, Candida has evolved several strategies to escape the host immune system. A detailed understanding of how C. tropicalis escapes the host immune attack is needed as it can help develop novel anti-fungal therapies. Secreted aspartyl proteinases (Saps) of C. albicans have been shown to be determinants of virulence and immune evasion. However, the immune evasion properties of C. tropicalis Saps have been poorly characterised. This study investigated the immune evasion properties of C. tropicalis secreted aspartic protease 1 (Sapt1).Sapt1 was recombinantly produced using a Kluyveromyces lactis yeast expression system. A range of complement proteins and immunogloublins were screened to test if Sapt1 had any proteolytic activity. Sapt1 efficiently cleaved human mannose-binding lectin (MBL) and collectin-11, which are the initiating molecules of the lectin pathway of the complement system, but not l-ficolin. In addition, Sapt1 cleaved DC-SIGN, the receptor on antigen presenting dendritic cells. Proteolysis was prominent in acidic condition (pH 5.2), a characteristic of aspartyl protease. No proteolytic activity was detected against complement proteins C1q, C3, C3b, IgG and IgA. In view of the ability of Sapt1 to cleave MBL and collectin-11, we found that Sapt1 could prevent activation of the complement lectin pathway. RT-qPCR analysis using three different C. tropicalis clinical isolates (oral, blood and peritoneal dialysis fluid) revealed relatively higher levels of mRNA expression of Sapt1 gene when compared to a reference strain; Sapt1 protein was found to be secreted by all the tested strains. Lectin pathway and its initiating components are crucial to provide front line defence against Candida infections. For the first time, we have shown that a Candida protease can proteolytically degrade the key initiating components of lectin pathway and inhibit complement activation. Findings from this study highlight the importance of exploring Sapt1 as a potential therapeutic target. We conclude that C. tropicalis secretes Sapt1 to target the complement lectin pathway, a key pattern recognition and clearance mechanism, for its survival and pathogenesis.

Serum Uromodulin and All-Cause Mortality in Peritoneal Dialysis Patients: A Chinese Cohort Study

Serum Uromodulin and All-Cause Mortality in Peritoneal Dialysis Patients: A Chinese Cohort Study

A graphene nanoplatelet-polydopamine molecularly imprinted biosensor for Ultratrace creatinine detection

Accurate and reliable analysis of creatinine is clinically important for the early detection and monitoring of patients with kidney disease. We report a novel graphene nanoplatelet (GNP)/polydopamine (PDA)-molecularly imprinted polymer (MIP) biosensor for the ultra-trace detection of creatinine in a range of body fluids. Dopamine hydrochloride (DA) monomers were polymerized using a simple one-pot method to form a thin PDA-MIP layer on the surface of GNP with high density of creatinine recognition sites. This novel surface-MIP strategy resulted in a record low limit-of-detection (LOD) of 2 × 10−2 pg/ml with a wide dynamic detection range between 1 × 10−1-1 × 109 pg/ml. The practical application of this GNP/PDA-MIP biosensor has been tested by measuring creatinine in human serum, urine, and peritoneal dialysis (PD) fluids. The average recovery rate was 93.7–109.2% with relative standard deviation (RSD) below 4.1% compared to measurements made using standard clinical laboratory methods. Our GNP/PDA-MIP biosensor holds high promise for further development as a rapid, accurate, point-of-care diagnostic platform for detecting and monitoring patients with kidney disease.

Steviol glycosides as an alternative osmotic agent for peritoneal dialysis fluid.

Background: Peritoneal dialysis (PD) is a renal replacement technique that requires repeated exposure of the peritoneum to hyperosmolar PD fluids (PDFs). Unfortunately, it promotes alterations of the peritoneal membrane (PM) that affects its functionality, including mesothelial-mesenchymal transition (MMT) of mesothelial cells (MCs), inflammation, angiogenesis, and fibrosis. Glucose is the most used osmotic agent, but it is known to be at least partially responsible, together with its degradation products (GDP), for those changes. Therefore, there is a need for more biocompatible osmotic agents to better maintain the PM. Herein we evaluated the biocompatibility of Steviol glycosides (SG)-based fluids. Methods: The ultrafiltration and transport capacities of SG-containing and glucose-based fluids were analyzed using artificial membranes and an in vivo mouse model, respectively. To investigate the biocompatibility of the fluids, Met-5A and human omental peritoneal MCs (HOMCs) were exposed in vitro to different types of glucose-based PDFs (conventional 4.25% glucose solution with high-GDP level and biocompatible 2.3% glucose solution with low-GDP level), SG-based fluids or treated with TGF-β1. Mice submitted to surgery of intraperitoneal catheter insertion were treated for 40 days with SG- or glucose-based fluids. Peritoneal tissues were collected to determine thickness, MMT, angiogenesis, as well as peritoneal washings to analyze inflammation. Results: Dialysis membrane experiments demonstrated that SG-based fluids at 1.5%, 1%, and 0.75% had a similar trend in weight gain, based on curve slope, as glucose-based fluids. Analyzing transport capacity in vivo, 1% and 0.75% SG-based fluid-exposed nephrectomized mice extracted a similar amount of urea as the glucose 2.3% group. In vitro, PDF with high-glucose (4.25%) and high-GDP content induced mesenchymal markers and angiogenic factors (Snail1, Fibronectin, VEGF-A, FGF-2) and downregulates the epithelial marker E-Cadherin. In contrast, exposition to low-glucose-based fluids with low-GDP content or SG-based fluids showed higher viability and had less MMT. In vivo, SG-based fluids preserved MC monolayer, induced less PM thickness, angiogenesis, leukocyte infiltration, inflammatory cytokines release, and MMT compared with glucose-based fluids. Conclusion: SG showed better biocompatibility as an osmotic agent than glucose in vitro and in vivo, therefore, it could alternatively substitute glucose in PDF.

Assessment of microbiological profile in peritoneal dialysis in patients with chronic kidney disease

Peritonitis is a serious complication of peritoneal dialysis (PD). It is the primary reason for hospitalization and switching over to hemodialysis. In the present study, we aimed to determine the microbiological profile of peritoneal dialysis fluid in patients with acute and chronic renal failure and identify these organisms' susceptibility patterns. This cross-sectional study was conducted in the Institute of Microbiology and Institute of Nephrology in the Rajiv Gandhi Government General Hospital from April 2016 to March 2017. A total of 100 patients who were &amp;#62;18 years of age, acute and chronic renal failure patients who underwent Peritoneal Dialysis and patients on Continuous and Intermittent Peritoneal Dialysis were included. The study population included 100 patients who satisfied the inclusion criteria. 63% were males, and 37% were females. The patients had a mean age of 44.15±13.89. 28 samples were culture positive, out of which 13 (46.4%) were Gram-negative, 10 (35.7%) were Gram-positive and 5 (17.9%) were Fungal isolates. Among them, the majority were Acinetobacter baumannii (20%) and Candida non-albicans (20%), followed by Staphylococcus aureus (16%), Coagulase-negative Staphylococcus (16%), Klebsiella oxytoca (8%), Klebsiella pneumoniae (4%), Enterococcus faecalis (4%).Peritoneal infections were more common in patients with longer duration of dialysis and diabetes mellitus. CAPD patients were having a higher risk of infections compared to IPD. As the number of infections associated with peritoneal dialysis rises, routine PD fluid microbiological analysis after the procedure will help improve patient care by using appropriate antibiotics as soon as possible before any major clinical problem arises.

Simultaneous determination and pharmacokinetics study of valsartan, sacubitril and its major metabolite in human plasma, urine and peritoneal dialysis fluid in patients with end-stage renal disease by UPLC–MS/MS

Sacubitril/valsartan was indicated for the treatment of heart failure and hypertension in patients with end-stage renal disease on peritoneal dialysis therapy. Herein, a rapid, sensitive and robust method based on ultra-liquid chromatography–tandem mass spectrometry (UPLC–MS/MS) was developed and validated for the determination of the concentrations of valsartan, sacubitril and its major metabolite LBQ657 in plasma, urine and peritoneal dialysis fluid. Samples were extracted from various biological fluids using protein precipitation. Extracts were subjected to UPLC–MS/MS with electrospray ionization in positive-ion mode. The developed method was fully validated over a concentration range of 8.00–8000 ng/mL for valsartan, 2.00–2000 ng/mL for sacubitril and 30.0–30,000 ng/mL for LBQ657 in plasma, 2.00–1000 ng/mL for valsartan, 1.00–500 ng/mL for sacubitril and 20.0–10000 ng/mL for LBQ657 in urine, and 0.200–100 ng/mL for valsartan, 0.0400–20.0 ng/mL for sacubitril and 2.00–1000 ng/mL for LBQ657 in peritoneal dialysis fluid. The intra- and inter-day precisions for all analytes in various matrices were less than 12.3%, and the intra- and inter-day accuracies results were all between 88.0% and 109.2%. All analytes were stable for at least 8 h at room temperature (25°C), five freeze–thaw cycles, and 37 days at −40°C and −80°C in plasma, urine and peritoneal dialysis fluid. In conclusion, this developed method is reliable, sensitive and specific for determining the concentrations of valsartan, sacubitril and LBQ657 in plasma, urine and peritoneal dialysis fluid. The assay was available to pilot the pharmacokinetics study of sacubitril/valsartan in patients with end-stage renal disease on peritoneal dialysis, and it could provide evidence that peritoneal dialysis had an effect on the clearance of sacubitril/valsartan.

Proteome-Wide Differential Effects of Peritoneal Dialysis Fluid Properties in an In Vitro Human Endothelial Cell Model.

To replace kidney function, peritoneal dialysis (PD) utilizes hyperosmotic PD fluids with specific physico-chemical properties. Their composition induces progressive damage of the peritoneum, leading to vasculopathies, decline of membrane function, and PD technique failure. Clinically used PD fluids differ in their composition but still remain bioincompatible. We mapped the molecular pathomechanisms in human endothelial cells induced by the different characteristics of widely used PD fluids by proteomics. Of 7894 identified proteins, 3871 were regulated at least by 1 and 49 by all tested PD fluids. The latter subset was enriched for cell junction-associated proteins. The different PD fluids individually perturbed proteins commonly related to cell stress, survival, and immune function pathways. Modeling two major bioincompatibility factors of PD fluids, acidosis, and glucose degradation products (GDPs) revealed distinct effects on endothelial cell function and regulation of cellular stress responses. Proteins and pathways most strongly affected were members of the oxidative stress response. Addition of the antioxidant and cytoprotective additive, alanyl-glutamine (AlaGln), to PD fluids led to upregulation of thioredoxin reductase-1, an antioxidant protein, potentially explaining the cytoprotective effect of AlaGln. In conclusion, we mapped out the molecular response of endothelial cells to PD fluids, and provided new evidence for their specific pathomechanisms, crucial for improvement of PD therapies.

A case of Moesziomyces antarcticus peritonitis in a patient undergoing peritoneal dialysis

BackgroundFungal peritonitis is a common and serious complication associated with peritoneal dialysis (PD), and it is often refractory to treatment.Case presentationA 70-year-old patient undergoing peritoneal dialysis was admitted to our hospital with fever and cloudy PD fluid. A diagnosis of yeast-like fungal peritonitis was made by examining the PD fluid. After starting intravenous caspofungin acetate, the PD catheter was removed. A fungal pathogen was isolated from the peritoneal fluid and identified as Cryptococcus sp. Based on the results of antifungal susceptibility testing, the treatment was changed to oral voriconazole and continued for 6 weeks. However, because of the discrepancy between the morphological findings and culture results, we performed genetic analysis, which uncovered Moesziomyces antarcticus. This patient was diagnosed with PD-related peritonitis caused by M. antarcticus and was successfully treated with voriconazole and removal of the PD catheter.ConclusionsReports of human infection by M. antarcticus are rare, and only two cases have been recognized. This may be the first case of infection detected in PD fluid.

A Culture-antibiogram of Sterile Body Fluids in a Tertiary Care Hospital at Central Delhi in Emergency Settings

Infection of sterile body fluids could be life-threatening. Monitoring of prevailing infectious agents and their resistance pattern through antibiogram helps in appropriate selection of antibiotics and strengthens antibiotic stewardship. The aim of this study is to isolate infective bacteria from sterile body fluids and to determine antibiotic resistant pattern of these isolates. This retrospective observational study was conducted through January 2018–December 2018 in a tertiary care hospital at central Delhi. Sterile body fluids were processed as per the standard laboratory procedures. Antibiotic susceptibility pattern was determined by Kirby Bauer’s disk diffusion method and interpreted as per Clinical and Laboratory Standards Institute guidelines. A total of 3,703 fluid samples were studied. Out of them 49.9% were ascitic fluid, 39.4% were pleural fluid, and 10.7% were others such as pericardial fluid, synovial fluid, bile, peritoneal dialysis fluid, and bronchoalveolar lavage fluid. Bacterial isolation rate was 12.93%. Predominant organisms were Escherichia coli (31%), Acinetobacter spp. (20%), Klebsiella spp. (16%), Staphylococcus aureus (11%), and Enterococcus spp. (10%). Isolation of vancomycin resistant enterococcus was 9.5% and Methicillin resistant S. aureus was 15.6%. Gram-negative isolates were 100% sensitive to colistin, followed by imipenem and gentamicin. Increased resistance shown toward third generation cephalosporins, amoxicillin + clavulinic acid, and piperacillin + tazobactum. Klebsiella spp. was the most resistant isolate, while Enterococcus spp. was the most resistant among gram positive isolates. Changing antimicrobial pattern poses challenge in treating infective agents. This culture antibiogram helps in appropriate selection of antibiotics in our setting and eventually decrease antibiotic resistant and patient’s morbidity and mortality.

Pharmacologic Inhibition of Histone Deacetylase 6 Prevents the Progression of Chlorhexidine Gluconate-Induced Peritoneal Fibrosis by Blockade of M2 Macrophage Polarization.

Peritoneal fibrosis contributes to ultrafiltration failure in peritoneal dialysis (PD) patients and thus restricts the wide application of PD in clinic. Recently we have demonstrated that histone deacetylase 6 (HDAC6) is critically implicated in high glucose peritoneal dialysis fluid (HG-PDF) induced peritoneal fibrosis, however, the precise mechanisms of HDAC6 in peritoneal fibrosis have not been elucidated. Here, we focused on the role and mechanisms of HDAC6 in chlorhexidine gluconate (CG) induced peritoneal fibrosis and discussed the mechanisms involved. We found Tubastatin A (TA), a selective inhibitor of HDAC6, significantly prevented the progression of peritoneal fibrosis, as characterized by reduction of epithelial-mesenchymal transition (EMT) and extracellular matrix (ECM) protein deposition. Inhibition of HDAC6 remarkably suppressed the expression of matrix metalloproteinases-2 (MMP2) and MMP-9. Administration of TA also increased the expression of acetylation Histone H3 and acetylation α-tubulin. Moreover, our results revealed that blockade of HDAC6 inhibited alternatively M2 macrophages polarization by suppressing the activation of TGF-β/Smad3, PI3K/AKT, and STAT3, STAT6 pathways. To give a better understanding of the mechanisms, we further established two cell injured models in Raw264.7 cells by using IL-4 and HG-PDF. Our in vitro experiments illustrated that both IL-4 and HG-PDF could induce M2 macrophage polarization, as demonstrated by upregulation of CD163 and Arginase-1. Inhibition of HDAC6 by TA significantly abrogated M2 macrophage polarization dose-dependently by suppressing TGF-β/Smad, IL4/STAT6, and PI3K/AKT signaling pathways. Collectively, our study revealed that blockade of HDAC6 by TA could suppress the progression of CG-induced peritoneal fibrosis by blockade of M2 macrophage polarization. Thus, HDAC6 may be a promising target in peritoneal fibrosis treatment.

A Clinically Relevant Murine Model of Peritoneal Fibrosis by Dialysate and Catheters.

Peritoneal fibrosis can occur in patients undergoing peritoneal dialysis (PD), and patients with severe peritoneal fibrosis have high morbidity and mortality. Peritonitis, high-glucose peritoneal dialysis fluid, and a long period of PD precipitate the onset of peritoneal fibrosis. An animal study of peritoneal fibrosis is needed due to the limitations of human and in vitro studies. However, most animal models do not mimic clinical conditions. To study peritoneal fibrosis, we developed a clinically relevant murine model by implanting a peritoneal catheter and injecting 2.5% high-glucose PD fluid plus 20 mM methylglyoxal (MGO) into the peritoneal cavity daily for 21 days. Implantation of the peritoneal catheter avoids peritoneal injury by needles and mimics clinical PD patients. Immunofluorescence staining showed that myofibroblasts accumulated in the fibrotic peritoneum. The experimental group had lower ultrafiltration volume and peritoneal membrane transport function (peritoneal equilibration test). In this article, we provide a detailed protocol of the model.

Enhanced Oxidative DNA-Damage in Peritoneal Dialysis Patients via the TXNIP/TRX Axis.

Peritoneal dialysis (PD) is an effective method of renal replacement therapy, providing a high level of patient autonomy. Nevertheless, the long-term use of PD is limited due to deleterious effects of PD fluids to the structure and function of the peritoneal membrane leading to loss of dialysis efficacy. PD patients show excessive oxidative stress compared to controls or chronic kidney disease (CKD) patients not on dialysis. Therefore, defense systems against detrimental events play a pivotal role in the integrity of the peritoneal membrane. The thioredoxin-interacting-protein (TXNIP)/thioredoxin (TRX) system also plays a major role in maintaining the redox homeostasis. We hypothesized that the upregulation of TXNIP negatively influences TRX activity, resulting in enhanced oxidative DNA-damage in PD patients. Therefore, we collected plasma samples and human peritoneal biopsies of healthy controls and PD patients as well. Using ELISA-analysis and immunohistochemistry, we showed that PD patients had elevated TXNIP levels compared to healthy controls. Furthermore, we demonstrated that PD patients had a reduced TRX activity, thereby leading to increased oxidative DNA-damage. Hence, targeting the TXNIP/TRX system as well as the use of oxidative stress scavengers could become promising therapeutic approaches potentially applicable in clinical practice in order to sustain and improve peritoneal membrane function.

MO711: Evaluation of an in Vitro Co-Culture Model for Studying Modulation of Cross-Talk between Endothelial and Mesothelial Cells by Cytoprotective Additives in Peritoneal Dialysis Fluids

Abstract BACKGROUND AND AIMS Peritoneal dialysis (PD) is a life-saving renal replacement therapy. The composition of all currently available peritoneal dialysis fluids (PDF) triggers morphological and functional changes in the peritoneal membrane. Periodic exposure leads to vasculopathy, hypervascularization and diabetes-like damage of vessels, eventually leading to failure of the technique. In vitro and in vivo studies have shown that cytoprotective additives [e.g. dipeptide alanyl-glutamine (AlaGln) or kinase inhibitor lithium chloride (LiCl)] to PDF reduce peritoneal damage in mesothelial (MC) and endothelial (EC) cells. However, it is yet unclear if potential cross talk between cells present in the peritoneal membrane mediates the observed modulation of PD-associated changes. Currently, there is no model system available to study relevant processes in these cells when in close proximity. Here, we aimed to develop a co-culture model for investigating cell-to-cell communication in a PD-relevant setting using cellular proteome and secretome analyses and investigate the effects of cytoprotective additives. METHOD For modelling the peritoneal membrane in vitro, MCs and ECs were co-cultured in transwell plates. MCs were grown in the upper compartment and primary microvascular ECs were grown in the lower compartment. PDF with or without additives, was added to the upper compartment to expose MCs directly, while the ECs below were kept in medium. Cellular proteome and secretome profiles were analysed for both cell types cultured individually or together by a quantitative mass spectrometry approach. Prior to analysis of the secretome, a combinatorial peptide ligand library coupled to beads was employed to enrich low abundant proteins and deplete high abundant proteins. RESULTS Proteome analysis revealed perturbation of major cellular processes including cytoskeleton reorganization, stress response, and regulation of cell death that characterize PDF cytotoxicity. Co-cultured cells yielded differently regulated pathways following PDF exposure compared to individual cultures. Several pathways relevant for PD, such as VEGF signaling in ECs or oxidative stress response in MCs were found only under co-culture conditions. Combined analysis of proteome and secretome showed specific ligand–receptor pairs expressed only under co-culture conditions regulating some of these pathways. Differentially regulated cellular and secreted proteins after PDF exposure were related to processes like angiogenesis, immune response, or and extracellular processes such as integrin signaling. Addition of 10 mM LiCl to PDF led to modulation of VEGF and oxidative stress response pathways in ECs as well as ligand–receptor pairs involved in these pathways, possibly explaining MC–EC crosstalk during PDF exposure of these cells and the cytoprotective effect of LiCl. CONCLUSION Harmful effects of PDF on MCs may also affect ECs, as demonstrated by our data. Interestingly, both cells type react differently when co-cultured compared to individual culture models, showing the importance of models that allow cell type interaction to mimic the in vivo conditions. We identified potential signaling axes between the cell types that could explain pathophysiological changes of the peritoneal membrane during PD treatment. We have also elucidated potential mechanisms by which the cytoprotective additive LiCl may modulate crosstalk between MCs and ECs to counteract adverse PDF effects in the local peritoneal environment. Linking local peritoneal damage with systemic vasculopathy through perturbations of PD-induced cellular crosstalk, may identify therapeutic targets to reduce the cardiovascular risk of PD patients and current limitations of the therapy.