Renal artery occlusion in a young woman - a tale of mysterious thrombosis.

BACKGROUND: Renal artery thrombosis is rare and limited reports exist in the young population. The most common aetiology is thromboembolic disease or abdominal trauma in this population and isolated occurrences are extremely rare. We present the case of an 18-year-old woman with spontaneous unilateral renal artery thrombosis and infarction for whom reperfusion was achieved through endovascular intervention. The aetiology of her thrombosis remains unclear and is under investigation with differential diagnoses being fibromuscular dysplasia, large and medium vessel vasculitis, and thromboembolic causes. OBJECTIVE: To demonstrate the value in attempting salvage of an ischaemic kidney in a young patient with an unexplained spontaneous renal thrombosis. METHOD: JM is an 18-year-old woman who presented to a large regional tertiary hospital with 3 days of right flank pain. She had no infective symptoms and no urinary or bowel changes before admission. She was not pregnant, and her only medication was the oral contraceptive pill commenced 3 months prior. A CT angiogram demonstrated right renal artery thrombosis with renal infarction. The kidney was deemed potentially salvageable, and ultrasound defined adequate vessel calibre to access for thrombectomy in the context of a negative coagulopathy screen. RESULTS: The patient underwent thrombolysis, thrombectomy and balloon angioplasty. Intraoperatively, a thin segment of distal stenosis was identified, and angiogram reperfusion was achieved with subsequent improvement in renal function. CONCLUSION: Renal artery thrombosis in young people is extremely rare and presents a diagnostic and management challenge requiring input from multiple teams including nephrology, rheumatology, paediatrics and vascular surgery. Systemic coagulopathy and vasculitis are differentials against anatomical aetiologies such as fibromuscular dysplasia. Our case adds to the limited literature regarding this in the young population. Renal artery thrombosis with occlusion in young people is very rare and is most often associated with a systemic coagulopathic disorder, such as antiphospholipid syndrome or structural pathology of the renal vasculature such as fibromuscular dysplasia. The work-up of a young female presenting with renal artery thrombosis without any previous medical history screens for a wide range of pathological processes.

Assessing left atrial size and pump function in ischemic stroke patients: Is cardiac MRI superior to transthoracic echocardiography?

OBJECTIVES: Current guidelines recommend transthoracic echocardiography (TTE) following an ischemic stroke as the primary technique to identify cardiac abnormalities associated with an increased risk of cerebral embolism. It is unclear whether cardiac magnetic resonance imaging (cMRI), a technique shown to provide increased imaging resolution, may also enhance the cardiac assessment of ischemic stroke patients. We compared cMRI with TTE in the evaluation of Left Atrial (LA) size and pump function in a cohort of 44 patients with ischemic stroke. MATERIALS AND METHODS: The biplane method was utilized to acquire LA diameters as well as area measurements in both TTE and cMRI. We calculated LA volume (LAV), LAV index (LAVI), LA Global Longitudinal Strain (GLS) and LA pump function. Results were compared using paired two sample for means t-test. Lin's concordance correlation coefficient (CCC) and Bland-Altman methods quantified the agreement of measurements obtained by TTE and cMRI. RESULTS: LAVI measurements by cMRI were significantly larger (34.97 v. 28.81; p = 0.001) than by TTE. The concordance correlation demonstrated only a weak agreement between LA size measured by cMRI and TTE (ρc = 0.397; p= 0.001, 95% CI 0.16 - 0.59), and the Bland-Altman plot demonstrated that LAVI measured by cMRI averaged 6.3 ml/m2 larger magnitude than those obtained by TTE. CONCLUSIONS: Using TTE alone leads to an underestimation of LA abnormalities important in the evaluation of ischemic stroke patients. Nearly one in every five ischemic stroke patients evaluated based on the current guidelines may have a missed potential source of cardiac embolism.

Surgical Episodes of Care for Price Transparency Using the Episode Grouper for Medicare.

OBJECTIVE: To explore the use of an episode grouper to more accurately identify the complete set of surgical services typically provided in a surgical episode of care and the corresponding range of prices, using colectomy for cancer as the example. BACKGROUND: Price transparency is an important policy issue that will require surgeons to better understand the components and cost of care. METHODS: This study uses the Episode Grouper for Medicare business logic to construct colectomy surgical episodes of care for cancer using Medicare claims data for the Boston Hospital Referral Region from 2012 to 2015. Descriptive statistics show the mean reimbursement based on patient severity and stage of surgery, along with the number of unique clinicians billing for care and the mix of services provided. RESULTS: The Episode Grouper for Medicare episode grouper identified 3182 colectomies in Boston between 2012 and 2015, with 1607 done for cancer. The mean Medicare allowed amount per case is $29,954 and varies from $26,605 to $36,850 as you move from low to high-severity cases. The intrafacility stage is the most expensive ($23,175 on average) compared with the pre ($780) and post ($6,479) facility stages. There is tremendous heterogeneity in the service mix. CONCLUSIONS: Episode groupers are a potentially valuable tool for identifying variations in service mix and teaming patterns that correlate with a total price. By looking at patient care holistically, stakeholders can identify opportunities for price transparency and care redesign that have heretofore been hidden.

Unraveling the complexity of amyloid polymorphism using gold nanoparticles and cryo-EM.

Increasing evidence suggests that amyloid polymorphism gives rise to different strains of amyloids with distinct toxicities and pathology-spreading properties. Validating this hypothesis is challenging due to a lack of tools and methods that allow for the direct characterization of amyloid polymorphism in hydrated and complex biological samples. Here, we report on the development of 11-mercapto-1-undecanesulfonate-coated gold nanoparticles (NPs) that efficiently label the edges of synthetic, recombinant, and native amyloid fibrils derived from different amyloidogenic proteins. We demonstrate that these NPs represent powerful tools for assessing amyloid morphological polymorphism, using cryogenic transmission electron microscopy (cryo-EM). The NPs allowed for the visualization of morphological features that are not directly observed using standard imaging techniques, including transmission electron microscopy with use of the negative stain or cryo-EM imaging. The use of these NPs to label native paired helical filaments (PHFs) from the postmortem brain of a patient with Alzheimer's disease, as well as amyloid fibrils extracted from the heart tissue of a patient suffering from systemic amyloid light-chain amyloidosis, revealed a high degree of homogeneity across the fibrils derived from human tissue in comparison with fibrils aggregated in vitro. These findings are consistent with, and strongly support, the emerging view that the physiologic milieu is a key determinant of amyloid fibril strains. Together, these advances should not only facilitate the profiling and characterization of amyloids for structural studies by cryo-EM, but also pave the way to elucidate the structural basis of amyloid strains and toxicity, and possibly the correlation between the pathological and clinical heterogeneity of amyloid diseases.

Incidence, Risk Factors and Outcome of COVID-19 Associated AKI- A Study from South India.

BACKGROUND: Acute Kidney Injury (AKI) is a dreaded complication of Covid-19 infection with high morbidity and mortality. Limited data exists on Indian experience. In a tertiary care hospital situated in South India, we analysed the incidence, clinical profile and outcomes of patients diagnosed with AKI due to COVID-19. METHODS: Retrospective data of Adult cases admitted with COVID-19 over a 8 month period from April - November 2020 was collected. Incidence, Demographics, Clinical profile, Management and Outcomes of COVID-19 associated AKI were analysed. Primary outcome was In- hospital mortality. Secondary outcomes were Dialysis Requirement and Renal Recovery. RESULTS: 52 (7%) out of a total 718 patients with COVID-19 developed AKI. Mean Age was 58 years (IQR 51-69) with a striking male predominance of 92%.(Male:Female -9.4:1) (P< 0.001). Co morbidities seen were Diabetes in 38 (73%) and Hypertension in 31(59%) and Coronary Artery Disease in 17(32%). Fever with myalgia was seen in 29 (46%), Respiratory symptoms in 31(59%), Oliguria in 26(50%) and Diarrhea in 2 (3%) patients. At admission, Hypoxemia and Hypotension were seen in 27 (51%) and 16 (30%) patients respectively. Urinalysis revealed > 2+ dipstick Proteinuria in 24 (46%) and Microscopic hematuria in 16(34%) patients. 25 (48%) were admitted in Stage 3 AKI with a mean S.Creatinine level of 4.4 + 3.4 mg/dl. CT chest showed > 50% lung involvement in 23 patients (44%). Mechanical ventilation was required in 20(38%). Hemodialysis was required in 12 (23%). The median duration of hospitalisation was 10 + 5 days. Primary Outcome of Mortality occurred in 44% of AKI cohort in comparison to 7% in Non AKI cohort (Relative Risk[RR]6.2; 95% Confidence Interval[CI], 4.1 to 9.4) (P= 0.001). Hypoxemia [RR,3.76;95% CI,1.4-9.5], Hypotension [RR 2.54; CI,1.5-4 ], Low Serum albumin [RR1.6;CI,1.1 - 2.3] and Requirement for mechanical ventilation [RR,11.3; CI, 2.9 - 23 ] were significant risk factors for mortality. All 5 patients who required both mechanical ventilation and Dialysis died. 31 patients (59%) were treated with Remdesivir therapy without survival benefit. Significantly higher C-Reactive Protein, Interleukin -6, D-Dimer levels and lower serum albumin levels were seen in those who died. Among the 29 patients who survived, the estimated GFR (e GFR) had recovered in 12 (41%) at discharge. After further followup of 4-6 weeks, the total number of patients who recovered renal function rose to 21 (72%). CONCLUSION: Mortality in COVID associated AKI stood at 44%. Multiple factors contributed to high mortality such as Severe disease with hypotension and extensive pulmonary involvement, High Neutrophil to Lymphocyte Ratio, Absolute Monocyte Count, inflammatory markers, d-Dimer and low serum albumin. It was encouraging to note that 72% of survivors recovered renal function by 4-6 weeks after discharge from hospital which means that it is worth the struggle to treat AKI in COVID-19.

Incidence and Impact of Healthcare-associated Infections on Patients Primarily Admitted with Sepsis and Non-sepsis Diagnoses.

Objectives: To compare the incidence of healthcare-associated infections (HAI) and their outcomes between patients admitted to the ICU with sepsis and those admitted with non-sepsis diagnoses. Materials and methods: We performed a single-center, prospective, observational study of ICU patients at a tertiary level medical-surgical unit from April 2018 to October 2018. All patients admitted to the ICU with a length of stay (LOS) > 48 hours were included. Baseline data including demographics, comorbidities, and severity of illness scores were collected. Index occurrence of HAI in all these patients was noted and data regarding organ support and patient outcomes were recorded. The incidence, complications, ICU LOS, and 30-day mortality of HAI were compared between the patients admitted to ICU originally with sepsis and non-sepsis diagnoses. Results: A total of 271 patients were evaluated in our study (N = 106 for the sepsis group and N = 165 for the non-sepsis group). No significant difference between the groups was found in the incidence of HAI (29.2% in sepsis group vs 24.4% in non-sepsis group; p = 0.07). Complications (acute kidney injury (AKI): 71 vs 45%; p = 0.01, shock: 81 vs 55%; p = 0.05, need for mechanical ventilation (MV): 30 vs 15%; p = 0.04) were more common in sepsis group compared to the non-sepsis group. The ICU LOS (12.2 ± 5.2 days vs 8.8 ± 2.05 days; p = 0.01) was significantly longer in the sepsis group. There was no significant difference in 30-day mortality between the groups (45 vs 25%; p = 0.07). Conclusion: The incidence of HAI seems to be similar between patients admitted with sepsis and non-sepsis diagnoses. However, patients admitted with sepsis develop higher rates of organ failure secondary to HAI and have a longer ICU LOS compared to patients admitted with non-sepsis diagnoses. The mortality rate of HAI did not differ between these two groups. How to cite this article: Chintamani A, Prakash B, Abraham BK, Kumar S, Ramakrishnan N, Venkataraman R. Incidence and Impact of Healthcare-associated Infections on Patients Primarily Admitted with Sepsis and Non-sepsis Diagnoses. Indian J Crit Care Med 2021;25(3): 292-295.

Practice Patterns and Outcome of Extracorporeal Membrane Oxygenation Therapy for Severe Acute Respiratory Distress Syndrome in Indian ICUs.

INTRODUCTION: Extracorporeal membrane oxygenation (ECMO) is increasingly used in managing patients with severe acute respiratory distress syndrome (ARDS). The aim of the study is to describe the practice of ECMO and evaluate the outcomes in patients with severe ARDS in Indian intensive care units (ICUs). METHODS: Data of 39 patients with severe ARDS managed with ECMO in two tertiary centers between 2012 and 2018 were retrospectively studied. RESULTS: The mean age of the patient was 44.6 ± 13.5 years and 51% were female. Primary ARDS due to viral pneumonia was the common indication for ECMO. Mean APACHE II and Murray scores were 32.3 ± 7.8 and 3.64 ± 0.21, respectively. Prone ventilation and/or inhaled nitric oxide were used in 69.3% of the patients prior to ECMO therapy. Among 39 patients, 38 patients were managed with venovenous ECMO and 1 patient was managed with venoarterial ECMO. Average ECMO duration was 9.4 ± 6.9 days. Among the 17 (43.5%) patients successfully weaned off ECMO, 15 (38.5%) survived to discharge home. The average ICU and hospital length of stay were 18.9 ± 15.5 and 20.6 ± 16.6 days, respectively. While, sepsis was the common complication noted in 19 (49%) patients, bleeding and thrombotic complications were also noted in six and two patients, respectively. CONCLUSION: In conclusion, ECMO support was used as rescue therapy in severe ARDS with a survival rate of 39%. Sepsis was the common complication of ECMO followed by bleeding and thrombosis. HOW TO CITE THIS ARTICLE: Mariappan R, Kumar M, Ramakrishnan N, Mani AK, Kumar S, Chandrasekaran V. Practice Patterns and Outcome of Extracorporeal Membrane Oxygenation Therapy for Severe Acute Respiratory Distress Syndrome in Indian ICUs. Indian J Crit Care Med 2021;25(11):1263-1268.

A simple, versatile and robust centrifugation-based filtration protocol for the isolation and quantification of α-synuclein monomers, oligomers and fibrils: Towards improving experimental reproducibility in α-synuclein research.

Increasing evidence suggests that the process of alpha-synuclein (α-syn) aggregation from monomers into amyloid fibrils and Lewy bodies, via oligomeric intermediates plays an essential role in the pathogenesis of different synucleinopathies, including Parkinson's disease (PD), multiple system atrophy and dementia with Lewy bodies (DLB). However, the nature of the toxic species and the mechanisms by which they contribute to neurotoxicity and disease progression remain elusive. Over the past two decades, significant efforts and resources have been invested in studies aimed at identifying and targeting toxic species along the pathway of α-syn fibrillization. Although this approach has helped to advance the field and provide insights into the biological properties and toxicity of different α-syn species, many of the fundamental questions regarding the role of α-syn aggregation in PD remain unanswered, and no therapeutic compounds targeting α-syn aggregates have passed clinical trials. Several factors have contributed to this slow progress, including the complexity of the aggregation pathways and the heterogeneity and dynamic nature of α-syn aggregates. In the majority of experiment, the α-syn samples used contain mixtures of α-syn species that exist in equilibrium and their ratio changes upon modifying experimental conditions. The failure to quantitatively account for the distribution of different α-syn species in different studies has contributed not only to experimental irreproducibility but also to misinterpretation of results and misdirection of valuable resources. Towards addressing these challenges and improving experimental reproducibility in Parkinson's research, we describe here a simple centrifugation-based filtration protocol for the isolation, quantification and assessment of the distribution of α-syn monomers, oligomers and fibrils, in heterogeneous α-syn samples of increasing complexity. The protocol is simple, does not require any special instrumentation and can be performed rapidly on multiple samples using small volumes. Here, we present and discuss several examples that illustrate the applications of this protocol and how it could contribute to improving the reproducibility of experiments aimed at elucidating the structural basis of α-syn aggregation, seeding activity, toxicity and pathology spreading. This protocol is applicable, with slight modifications, to other amyloid-forming proteins.

How specific are the conformation-specific α-synuclein antibodies? Characterization and validation of 16 α-synuclein conformation-specific antibodies using well-characterized preparations of α-synuclein monomers, fibrils and oligomers with distinct structures and morphology.

Increasing evidence suggests that alpha-synuclein (α-syn) oligomers are obligate intermediates in the pathway involved in α-syn fibrillization and Lewy body (LB) formation, and may also accumulate within LBs in Parkinson's disease (PD) and other synucleinopathies. Therefore, the development of tools and methods to detect and quantify α-syn oligomers has become increasingly crucial for mechanistic studies to understand their role in PD, and to develop new diagnostic methods and therapies for PD and other synucleinopathies. The majority of these tools and methods rely primarily on the use of aggregation state-specific or conformation-specific antibodies. Given the impact of the data and knowledge generated using these antibodies on shaping the foundation and directions of α-syn and PD research, it is crucial that these antibodies are thoroughly characterized, and their specificity or ability to capture diverse α-syn species is tested and validated. Herein, we describe an antibody characterization and validation pipeline that allows a systematic investigation of the specificity of α-syn antibodies using well-defined and well-characterized preparations of various α-syn species, including monomers, fibrils, and different oligomer preparations that are characterized by distinct morphological, chemical and secondary structure properties. This pipeline was used to characterize 18 α-syn antibodies, 16 of which have been reported as conformation- or oligomer-specific antibodies, using an array of techniques, including immunoblot analysis (slot blot and Western blot), a digital ELISA assay using single molecule array technology and surface plasmon resonance. Our results show that i) none of the antibodies tested are specific for one particular type of α-syn species, including monomers, oligomers or fibrils; ii) all antibodies that were reported to be oligomer-specific also recognized fibrillar α-syn; and iii) a few antibodies showed high specificity for oligomers and fibrils but did not bind to monomers. These findings suggest that the great majority of α-syn aggregate-specific antibodies do not differentiate between oligomers and fibrils, thus highlighting the importance of exercising caution when interpreting results obtained using these antibodies. Our results also underscore the critical importance of the characterization and validation of antibodies before their use in mechanistic studies and as diagnostic tools or therapeutic agents. This will not only improve the quality and reproducibility of research and reduce costs but will also reduce the number of therapeutic antibody failures in the clinic.

Impact of Fluoridated and Non-fluoridated Mouth Rinses on Frictional Resistance between Orthodontic Archwire and Bracket: A Comparative Study.

AIM AND OBJECTIVE: The present study aimed to assess the effect of fluoridated and nonfluoridated mouth ri nses on resistance to friction between orthodontic bracket and archwire. MATERIALS AND METHODS: This study comprises 60 premolar stainless steel (SS) brackets with 0.022 inches slot size. The 0.019 × 0.025 dimensions SS archwires were cut into 5 cm long specimens. They were grouped into three main categories, group I: artificial saliva (control solution), group II: Aloe Dent mouthwash (ALO), and group III: 0.05% sodium fluoride mouthwash. The specimens from each group were either immersed in the test solution or in the control solution for 10 hours. Later, the specimens were transferred to an incubator maintained at 37°C. Post 10 hours, the specimens were immersed for 30 minutes in distilled water. A scanning electron microscope was used to study the surface morphology and a universal testing machine was used to measure the frictional resistance. RESULTS: The distribution of normality for three study groups' recorded data was checked using Shapiro-Wilk test. The highest frictional resistance (1.94 ± 0.02) was demonstrated by specimens immersed in 0.05% sodium fluoride mouthwash than those immersed in Aloe Dent mouthwash (1.28 ± 0.66) and artificial saliva (1.10 ± 0.32). The difference found between the groups by an analysis of covariance was statistically significant. The highest surface roughness (22.30 ± 0.12) was revealed by specimens immersed in 0.05% sodium fluoride mouthwash than those immersed in Aloe Dent mouthwash (18.28 ± 0.26) and artificial saliva (15.86 ± 0.42). A statistically significant difference between the groups was shown by an analysis of covariance. CONCLUSION: After considering the drawbacks of this study, we conclude that specimens immersed in Aloe Dent mouthwash demonstrated less frictional resistance and surface roughness when compared to those immersed in 0.05% sodium fluoride mouthwash. CLINICAL SIGNIFICANCE: During sliding mechanism, the frictional resistance between orthodontic archwire and brackets imposes problems, such as lessening the applied force and movement of tooth, and also results in anchorage loss. So, orthodontists should always take care while prescribing mouthwashes to reduce their effects on the friction.

Multimodality detection of multiple left ventricular diverticula: A case report and brief review of the literature.

Left ventricular diverticula (LVD) are rare congenital anomalies usually detected incidentally in the adult population. Most commonly, they are found as a single left ventricular diverticulum in association with other congenital abnormalities but multiple LVD are exceedingly rare. We are describing a patient who was found to have multiple LVD on multimodality imaging studies. He had presented with a sudden cardiac arrest attributed to a combination of alcohol intoxication and QT interval prolongation from hypokalemia and antidepressant medications. The patient was managed conservatively and discharged with an implantable loop recorder for detecting any occult arrhythmias.

Artificial Intelligence and Deep Learning: The Future of Medicine and Medical Practice.

ABSTRACT: Artificial Intelligence (AI) and access to "Big Data" together with the evolving techniques in biotechnology will change the medical practice a big way. Many diseases such as type II diabetes will no longer be considered as a single disease. Many familiar cancers such as cancer of liver or pancreas will have hundreds of subtypes whose management will be very different. The way we think about diseases will change. It will no longer be possible for clinicians to make a diagnosis, remember the names of diseases, the names of drugs or management protocols without the help of computers. As computer intelligence becomes more important than human intelligence in deciding diagnosis and treatment there will be a paradigm in the role of doctors. Internet, computers and social media will become more important than individuals in decision making. As a result, medicine will go more and more egalitarian ("wiki") with increasing community participation in health decision making and management. A socialistic pattern will evolve over time globally as an adaptive reaction to the pressures put by artificial intelligence. This is because the individual differences in knowledge or intellect between human beings will become less apparent compared to the super powers of artificial intelligence. Qualities which are unique for humans such as compassion, empathy and emotional care will decide the professional success of future physicians even more than today. Today we are using artificial intelligence in diagnosis and prediction to help clinicians. Clinical algorithms and human experience cannot be replaced by machines. It will take many years to completely merge or replace humans with machines. However, we need to modify our medical education system in order to prepare the medical community and sensitize the society well in advance for a smooth transition.

Comparative Evaluation of Hydrophobic and Hydrophilic Resin-based Sealants: A Clinical Study.

AIM: The purpose of this study is to clinically evaluate and compare the retention and evidence of caries of three fissure sealants. MATERIALS AND METHODS: A total of 150 children, between 7 and 13 years of age, with fully erupted permanent molars, had sealants placed using a full-mouth design. Sealant retention was evaluated at 1, 3, 6, 9, and 12 months later. Teeth were evaluated for retention and evidence of caries using Simonsen's criteria and results were subjected to statistical analysis using the Chi-square test. RESULTS: At 1-year examination, in teeth sealed with Clinpro: (a) 8% were completely retained, (b) 74.4% were partially lost, and (c) 8.5% were completely lost; with Embrace Wetbond: (a) none of the sealants were completely retained, (b) 13.1% were partially lost, and (c) 59.1% were completely lost; with Champ: (a) 1% were completely retained, (b) 71.4% were partially lost, and (c) 10.9% were completely lost. All the three sealants showed evidence of caries from 9 months. CONCLUSION: The retention of hydrophobic (Clinpro) sealant was superior to hydrophilic (Embrace Wetbond and Champ) sealants. The evidence of caries was less in the hydrophobic sealant group when compared with the hydrophilic sealant groups. There was no statistical difference in retention and evidence of caries between maxillary and mandibular teeth for all the three sealant groups. CLINICAL SIGNIFICANCE: Sealants prevent the occurrence of caries in the majority of children. Though hydrophobic sealants appear to be more successful, hydrophilic sealants too may provide promising results in the near future.

TRAF3IP2 mediates TWEAK/TWEAKR-induced pro-fibrotic responses in cultured cardiac fibroblasts and the heart.

Persistent inflammation promotes development and progression of heart failure (HF). TWEAK (TNF-Related WEAK Inducer Of Apoptosis), a NF-κB- and/or AP-1-responsive proinflammatory cytokine that signals via TWEAK receptor (TWEAKR), is expressed at high levels in human and preclinical models of HF. Since the adapter molecule TRAF3IP2 (TRAF3 Interacting Protein 2) is an upstream regulator of various proinflammatory pathways, including those activated by NF-κB and AP-1, we hypothesized that targeting TRAF3IP2 inhibits TWEAK-induced proinflammatory and pro-fibrotic responses in vitro and in vivo. Consistent with the hypothesis, forced expression of TRAF3IP2 upregulated TWEAK and its receptor expression in cultured adult mouse cardiac fibroblasts (CF). Further, exogenous TWEAK upregulated TRAF3IP2 expression in a time- and dose-dependent manner, suggesting a positive-feedback regulation of TRAF3IP2 and TWEAK. TWEAK also promoted TRAF3IP2 nuclear translocation. Confirming its critical role in TWEAK signaling, silencing TRAF3IP2 inhibited TWEAK autoregulation, TWEAKR upregulation, p38 MAPK, NF-κB and AP-1 activation, inflammatory cytokine expression, MMP and TIMP1 activation, collagen expression and secretion, and importantly, proliferation and migration. Recapitulating these in vitro results, continuous infusion of TWEAK for 7 days increased systolic blood pressure (SBP), upregulated TRAF3IP2 expression, activated p38 MAPK, NF-κB and AP-1, induced the expression of multiple proinflammatory and pro-fibrotic mediators, and interstitial fibrosis in hearts of wild type mice. These proinflammatory and pro-fibrotic changes occurred in conjunction with myocardial hypertrophy and contractile dysfunction. Importantly, genetic ablation of TRAF3IP2 inhibited these TWEAK-induced adverse cardiac changes independent of increases in SBP, indicating that TRAF3IP2 plays a causal role, and thus a therapeutic target, in chronic inflammatory and fibro-proliferative diseases.

Efficacy of Novel Antistaphylococcal Ectolysin P128 in a Rat Model of Methicillin-Resistant Staphylococcus aureus Bacteremia.

Staphylococcus aureus causes systemic infections with high morbidity and mortality, and the emergence of drug-resistant strains is a rapidly growing clinical concern. Novel therapeutic agents are required to tackle S. aureus infections. P128 is a bacteriophage-derived chimeric ectolysin with potent and rapid bactericidal activity against S. aureus In the present study, the efficacy of P128 was evaluated in a newly developed rat model of S. aureus bacteremia. Prior to in vivo testing, P128 was shown to be stable in whole blood by incubation in rat blood for up to 6 h and testing its bactericidal activity against the methicillin-resistant S. aureus isolate USA300. Rats succumbed to intravenous challenge with 109 CFU of S. aureus USA300, resulting in 80 to 100% mortality by day 14. Evaluation of the bacterial load in various organs at 96 h postinfection revealed high bacterial counts in the kidney, and this correlated with the presence of renal abscesses. Treatment of infected animals with P128 either by intravenous bolus administration via tail vein or by 1-h infusion via the jugular vein at 2 h postinfection resulted in the dose-dependent survival of rats. P128 treatment also resulted in very few or no abscesses in the kidneys. These data show that P128 is stable in the physiological milieu and that intravenous treatment with P128 is highly effective in rescuing rats from S. aureus bacteremia. P128 can be a novel therapeutic option for treatment of S. aureus systemic infections.

Efficacy of chimeric ectolysin P128 in drug-resistant Staphylococcus aureus bacteraemia in mice.

Objectives: P128 is a recombinant chimeric ectolysin with potent antistaphylococcal activity. P128 was evaluated as monotherapy and in combination with two standard-of-care (SoC) antibiotics, vancomycin and daptomycin, in mouse models of Staphylococcus aureus bacteraemia. Methods: Healthy BALB/c mice were challenged (intraperitoneally) with 109 cfu of MRSA strain COL or USA300 and treated with a single dose of P128 (0.2-10 mg/kg). Drug synergy was tested using a single dose of P128 (0.2 or 2.5 mg/kg) along with sub-therapeutic dose levels of vancomycin (27.5 or 55 mg/kg) or daptomycin (12.5 mg/kg). Bacterial load was checked in peritoneal fluid and in blood, at different time intervals. Synergy against drug-resistant strains was tested using the P128/vancomycin combination against vancomycin-resistant S. aureus (VRSA). Results: In MRSA bacteraemia, P128, vancomycin and daptomycin monotherapy resulted in 31%, 46% and 46% survival, respectively. The P128/vancomycin and P128/daptomycin combinations afforded increased survival of 85% and 88%, respectively. P128 showed a rapid bactericidal effect with a reduction of cfu in both the peritoneal fluid and the blood within 1 h. In VRSA bacteraemia, a mouse-equivalent therapeutic dose of vancomycin (110 mg/kg) failed to rescue animals. P128 (1-20 mg/kg) as monotherapy resulted in dose-dependent efficacy. Survival (37%) with 2.5 mg/kg P128 increased to 63% with the P128/vancomycin combination. Conclusions: P128 exerted a rapid bactericidal effect in vivo and rescued animals from fatal invasive MRSA and VRSA infections. P128/SoC antibiotic combinations exerted a synergistic effect. P128 restored the susceptibility of VRSA to vancomycin. P128 is a novel, potent therapeutic agent for antibiotic-resistant, systemic S. aureus infections.

A novel strategy to produce high level and high purity of bioactive IL15 fusion proteins from mammalian cells.

IL15, a member of the common γ chain receptor (γc) cytokine family, is gaining attention in recent years as one of the most promising anti-tumor agents. IL15 regulates T cell activation and proliferation, promotes the survival of CD8+ CD44hi memory T cells and is also essential for NK cell expansion and development. Despite the attraction of developing IL15 as an anti-cancer agent, production of recombinant IL15 has proven to be difficult due to the stringent control of IL15 expression at the transcriptional, translational and the post-translational levels. Furthermore, the bioactivity of IL15 fused to an extra functional domain that is isolated from mammalian cells is generally inferior to recombinant IL15 produced by E. coli. In this study, we report that Lysine 86 in IL15 is responsible for the instability in mammalian cells when its C-terminus is fused to the albumin binding scFv (IL15-A10m3). We demonstrate that K86A or K86R mutants increased the expression of the fusion protein from HEK293 cells. When the wild type IL15 is used for the fusion, no recombinant IL15 fusion was detected in the culture media. Additionally, we determined that the residue 112 in IL15 is critical for the bioactivity of IL15-A10m3. Examination of single and double mutants provides a better understanding of how IL15 engages with its receptor complex to achieve full signaling capacity. The results of our experiments were successfully applied to scale up production to levels up to 50 mg/L and >10 mg/L of >95% pure monomeric recombinant fusion proteins after a 2-step purification from culture media. More importantly, the recombinant fusion protein produced is fully active in stimulating T cell proliferation, when compared to the recombinant wild type IL15.

Donor CD163 and nestin-positive cells predict graft function in living donor liver transplant.

AIMS: To evaluate the effect of donor liver resident mesenchymal cells, M2 macrophages on liver graft outcome after living donor liver transplantation. MATERIALS AND METHODS: Seventy donor biopsies were included in the study. Outcomes at day 3, 7, 30, and 180 postliver transplantation were assessed. Mesenchymal stem cells and M2 macrophages in donor liver biopsies were evaluated. RESULTS: Mean age of recipients was 40.9 ± 13.6 years. Sex mismatched transplants were 44 (M→F = 9; F→M = 35). On area under receiver operative curve analysis, donor biopsy (DB) nestin ≥3 and CD 163 ≥ 32/200x at day 3; CD163 ≥ 32 at day 7; CD 163 > 32, pRBC of <6.5 units at day 30, and DB nestin ≥3, CD 163 ≥ 32 and pRBC<6.5 units at day 180 predicted adequate graft functions. On multivariate analysis, higher DB nestin (P = .009) and lower cryoprecipitate (P = .009) usage at day 3, higher DB CD163 (P = .006) at day 7, higher DB CD163 (P = .018) and reduced transfusion of packed cell (pRBC) (P = .014) at day 30 and higher DB nestin (P = .011), higher CD163 (P = .009), and reduced pRBC (P = .045) at day 180 were the predictors of better outcome. CONCLUSIONS: Donor liver biopsy nestin+ and CD163+ can predict early graft outcome in living donor liver transplantation.

Thermoluminescence from Cu Doped Lithium Tetraborate Irradiated with X-ray and γ Using 137Cs Radioactive Source.

Lithium tetraborate (Li2B4O7) pellets prepared by using water/solution assisted method were synthesized and characterized. Copper was used as doping material in order to enhance the Li2B4O7 thermoluminescent properties. For synthesis heating temperature parameters were defined at 750 °C for 2 hr, followed by 150 °C for another 2 hr. The materials were produced at five different Cu concentrations: 0.02, 0.04, 0.06, 0.08, and 0.1 wt%. The luminescent and morphological characterizations were performed by X-ray diffraction (XRD), Scanning electron microscope (SEM), Photoluminescence (PL), and Ultraviolet-visible spectroscopy (UV-Vis). XRD and SEM analysis of intrinsic and doped materials confirmed the obtained Li2B4O7 structure and show its morphology. XRD patterns of the Li2B4O7 matched a tetragonal crystal structure. Crystals of Li2B4O7 of an average size of 50 nm were obtained. The presence of the copper dopant was confirmed in crystals of Li2B4O7:Cu by SEM-EDS (energy dispersive spectroscopy X-ray). The emission spectrum of Cu doped Li2B4O7 showed a prominent peak at 367 nm, while the main UV-Vis absorption was observed from 240 nm to 300 nm due to Cu+ ion 3d10 → 3d9 4s transitions. The thermoluminescent (TL) response was studied for both γ radiation and X-ray. A 661.7 keV γ radiation using a 137Cs source at doses of 50, 100, 200, 300, 400 and 500 mGy was applied to Li2B4O7:Cu (0.1 wt%) pellets. An X-ray source was used at doses of 600, 800 and 1000 mGy to irradiate pellets of Li2B4O7:Cu (0.02, 0.04, 0.06, 0.08 and 0.1 wt%). A linear TL response was observed for both X-ray and γ radiation. The kinetic parameters were calculated using the peak shape method for the Li2B4O7:Cu (0.1 wt%).

Attenuation of Metabolic Syndrome by EPA/DHA Ethyl Esters in Testosterone-Deficient Obese Rats.

Inducing testosterone deficiency, as the standard treatment of prostate cancer, may cause metabolic disorders including insulin resistance, dyslipidemia, central obesity, cardiovascular diseases, and type 2 diabetes. This study measured responses to testosterone deficiency in high-carbohydrate, high-fat (H) diet-fed rats. We then tested whether eicosapentaenoic acid (EPA)/docosahexaenoic acid (DHA) ethyl esters (Omacor) reversed these metabolic changes. Male Wistar rats (8⁻9 weeks old) were divided into eight groups with four groups fed corn starch and four groups fed H diet. For each diet, one group received diet only; one group was orchidectomized; one group was given leuprolide (gonadotrophin-releasing hormone agonist, 2 mg/kg every 4th week); and the last group was treated with leuprolide and their diet was supplemented with 3% Omacor for the last eight weeks. The protocol was for 16 weeks. Leuprolide worsened metabolic syndrome symptoms and cardiovascular function, and orchidectomy produced greater responses. In H fed leuprolide-treated rats, Omacor decreased systolic blood pressure and left ventricular diastolic stiffness, reduced infiltration of inflammatory cells and collagen deposition in the heart, and reduced lipid accumulation and inflammatory cell infiltration without improving liver damage. These results suggest that Omacor has potential to attenuate metabolic complications in prostate cancer patients with induced testosterone deprivation.