Histopathological Severity Score Research Articles

Postmortem chest computed tomography in COVID-19: A minimally invasive autopsy method

ObjectivesPerforming autopsies in a pandemic scenario is challenging, as the need to understand pathophysiology must be balanced with the contamination risk. A minimally invasive autopsy might be a solution. We present a model that combines radiology and pathology to evaluate postmortem CT lung findings and their correlation with histopathology. MethodsTwenty-nine patients with fatal COVID-19 underwent postmortem chest CT, and multiple lung tissue samples were collected. The chest CT scans were analyzed and quantified according to lung involvement in five categories: normal, ground-glass opacities, crazy-paving, small consolidations, and large or lobar consolidations. The lung tissue samples were examined and quantified in three categories: normal lung, exudative diffuse alveolar damage (DAD), and fibroproliferative DAD. A linear index was used to estimate the global severity of involvement by CT and histopathological analysis. ResultsThere was a positive correlation between patient mean CT and histopathological severity score indexes - Pearson correlation coefficient (R) = 0.66 (p = 0.0078). When analyzing the mean lung involvement percentage of each finding, positive correlations were found between the normal lung percentage between postmortem CT and histopathology (R=0.65, p = 0.0082), as well as between ground-glass opacities in postmortem CT and normal lungs in histopathology (R=0.65, p = 0.0086), but negative correlations were observed between ground-glass opacities extension and exudative diffuse alveolar damage in histological slides (R=−0.68, p = 0.005). Additionally, it was found is a trend toward a decrease in the percentage of normal lung tissue on the histological slides as the percentage of consolidations in postmortem CT scans increased (R =−0.51, p = 0.055). The analysis of the other correlations between the percentage of each finding did not show any significant correlation or correlation trends (p ≥ 0.10). ConclusionsA minimally invasive autopsy is valid. As the severity of involvement is increased in CT, more advanced disease is seen on histopathology. However, we cannot state that one specific radiological category represents a specific pathological correspondent. Ground-glass opacities, in the postmortem stage, must be interpreted with caution, as expiratory lungs may overestimate disease.

Fungicides from rice cultivation (tebuconazole and azoxystrobin) alters biochemical and histological markers of hammertoad tadpoles (Boanafaber).

Tebuconazole (TBZ) and azoxystrobin (AZX) are fungicides frequently used in rice cultivation. Despite protecting crops against fungal diseases, these compounds can contaminate the natural environments close to the crops, exerting negative effects on non-target organisms, the present study aimed to characterize the contamination by fungicides of a river that flows in an area dominated by rice cultivation in the north of the state of Santa Catarina, SC, Brazil. Concentrations of TBZ and AZX found in the field were used to evaluate their negative effects on development, biochemical biomarkers and histopatology of the liver of a native tadpole species, the hammerfrog (Boana faber). Tadpoles were exposed for 16 days to the lowest (1.20 μg/L) and highest (2.60 μg/L) concentration of TBZ, lowest (0.70 μg/L) and highest (1.60 μg/L) concentration of AZX, and the mix of both fungicides at lowest and highest concentration of each found in field analyses. Exposure to the lower TBZ concentration and both concentrations of the Mix accelerated the development of tadpoles. AZX caused an increase in the activities of glutathione S-transferase (GST), carboxylesterase (CbE) and glucose-6-phosphate dehydrogenase (G6PDH) in the liver, an increase in the levels of protein carbonyls (PC) in the liver and an increase in the activity of acetylcholinesterase (AChE) in muscle of tadpoles. TBZ, on the other hand, generated an increase in GST, G6PDH, PC and histopathological severity scores in liver and in muscle AChE activity. The effects were more intense in the groups exposed to the Mix of contaminants. No treatment altered brain AChE. The data showed that the fungicides from in rice cultivation found in natural aquatic environments around the crops pose risks to the health of the animals, compromising their metabolism and development.

Association between the severity of histopathological lesions and Mycobacterium avium subspecies paratuberculosis (MAP) molecular diversity in cattle in southern Chile.

The objective was to evaluate the association between the severity of histopathological lesions caused by Mycobacterium avium subspecies paratuberculosis (MAP) infection and the molecular diversity of this pathogen. Blood, ileum, and mesenteric lymph node samples were collected at slaughter, from 1,352 adult cattle [source population 1 (SP1)]. In addition, 42 dairy herds (n = 4,963 cows) were followed for 2 years, and samples from compatible paratuberculosis clinical cases [source population 2 (SP2)] were collected. MAP infection was confirmed using an ELISA test, liquid media culture, and PCR. Isolates were genotyped using five MIRU-VNTR markers. Tissues from confirmed samples were subjected to a histopathological examination. A histopathological severity score (HSS) system was developed and used to grade (0 to 5) the magnitude of lesions caused by MAP. In general, the HSS system assesses the number of foci and degree of macrophage infiltration, together with the presence of multinucleated giant cells (MGCs) and acid-fast bacilli (AFB), in addition to the fusion of the intestinal villi and hyperplasia of the crypts. Despite the large sampling effort, only 79 MAP isolates were successfully genotyped, where 19 different haplotypes were described. A mixed-effect Poisson regression model was used to assess the relationship between haplotypes and HSS values. The model was controlled by animal age, and the farm was used as a random effect. Haplotypes were grouped based on their relative frequency: the most frequent haplotype (group i, 49.4%), the second most frequent haplotype (group ii, 12.7%), and all other haplotypes (group iii, 37.9%). Model outputs indicated that group i had significantly higher HSS values than group iii. In addition, group i was also associated with higher optical density (OD) values of the ELISA test. These results support the existence of differences in pathogenicity between MAP haplotypes. However, results were based on a relatively small sample size; thus, these should be taken with caution. Despite this, study findings suggest that haplotypes would be associated with differences in disease progression, where the dominant haplotype tends to generate more severe lesions, which could be linked to a greater shed of MAP cells than non-dominant haplotypes, increasing their chances of transmission.

Rejection and graft outcomes in kidney transplant recipients with and without angiotensin II receptor type 1 antibodies

AimAngiotensin II type 1 receptor antibody (AT1R Ab) is a non-Human Leucocyte Antigen (HLA) antibody that is maybe associated with early severe kidney transplant rejection and worse graft outcomes. This study aimed to assess the association between AT1R Ab and kidney transplant rejection and graft outcomes. MethodsWe performed a retrospective analysis of all adult kidney transplant recipients in an Australian centre who had an AT1R Ab test between 1 January 2015 to 30 June 2020. AT1R Ab positive patients were compared to AT1R Ab negative patients. Primary outcomes were rejection risk, type and histopathological severity scores. Secondary outcomes were 8-week graft function and graft loss. ResultsOf 965 kidney transplants that were performed during the study period, 73 patients had AT1R Ab tested; 16 (22%) were positive and 57(78%) were negative. Positive patients were on average younger and had higher level of donor-specific HLA antibodies. Rejection occurred in 13 (81%) positive patients and 41 (72%) negative patients (P = 0.45). No significant differences in rejection type or severity were found. HLA mismatch and peak panel reactive antibody ≥80%, but not AT1R Ab, independently predicted rejection. Average (132 vs. 177 mmol/L, P = 0.302) and graft loss were not significantly different between groups. ConclusionThe study found no evidence that AT1R Ab is associated with rejection type, severity or worse graft function. Future studies should assess its relationship with graft outcomes to help complement immunological risk assessment and potentially provide therapeutic options to alter outcomes.

The severity of chronic histiocytic intervillositis is associated with gestational age and fetal weight

IntroductionChronic histiocytic intervillositis (CHI) is a rare histopathological lesion in the placenta that is associated with poor reproductive outcomes. The intervillous infiltrate consists mostly of maternal mononuclear cells and fibrin depositions, which are both indicators for the severity of the intervillous infiltrate. The severity of the intervillous infiltrate as well as the clinical outcomes of pregnancy differ between cases. Our objective is to determine the relation between the severity of the intervillous infiltrate and the clinical outcomes of CHI. MethodsCases of CHI were semi-quantitatively graded based on histopathological severity scores. Hereto, CD68 positive mononuclear cells were quantified, fibrin depositions visualized by both a PTAH stain and an immuohistochemical staining, and placental dysfunction was assessed via thrombomodulin staining. ResultsThis study included 36 women with CHI. A higher CD68 score was significantly associated with a lower birthweight. Loss of placental thrombomodulin was associated with lower gestational age, lower birthweight, and a lower placenta weight. The combined severity score based on CD68 and PTAH was significantly associated with fetal growth restriction, and the joint score of CD68 and fibrin was associated with birthweight and placental weight. DiscussionMore severe intervillous infiltrates in CHI placentas is associated with a lower birth weight and placental weight. Furthermore, this study proposes thrombomodulin as a possible new severity marker of placental damage. More research is needed to better understand the pathophysiology of CHI.

Hepatic and blood alterations in Lithobates catesbeianus tadpoles exposed to sulfamethoxazole and oxytetracycline

In this study the effects of environmentally realistic concentrations of the antibiotics sulfamethoxazole (SMX) and oxytetracyclyne (OTC) on Lithobates catesbeianus tadpoles were evaluated, through the analyzes of the frequencies of micronucleus and nuclear abnormalities in erythrocytes, alterations in leucocytes, liver histopathology, and changes in hepatic esterase activities and oxidative stress biomarkers. The animals were exposed for 16 days at concentrations of 0 (control), 20, 90 and 460 ng L−1. No significant difference was found in the frequencies of micronucleus and nuclear abnormalities. The two highest concentrations of SMX and all concentrations of OTC caused a significant increase in the number of lymphocytes. A significant decrease in the number of neutrophils compared to the control group was observed for all concentrations tested of both antibiotics. Also, decrease in the activity of glutathione S-transferase and high histopathological severity scores, indicating liver damage, were found in tadpoles exposed to the two highest concentrations of SMX and all concentrations of OTC. The main changes in the liver histopathology were the presence of inflammatory infiltrate, melanomacrophages, vascular congestion, blood cells and eosinophils. Esterase activities were unchanged. Indeed, the two highest concentrations of OTC caused a reduction in the activities of superoxide dismutase and glucose 6-phosphate dehydrogenase, while the highest concentration inhibited the activity of glutathione peroxidase and increased protein carbonyl levels. These results evidences that environmentally realistic concentrations of SMX and OTC in aquatic environments are capable to significantly disrupt tadpoles’ physiology, possibly affecting negatively their survival rate in natural environments.

Relationship between Serum Protein Electrophoresis, Endoscopic and Histopathological Scores in 99 Cats with Chronic Enteropathy.

Simple SummarySerum protein electrophoresis (SPE) is a laboratory test used to separate different protein fractions. It is used to investigate animals with hyperglobulinemia and to distinguish between monoclonal and polyclonal gammopathies. SPE can also highlight acute or chronic inflammation patterns which are useful in disease monitoring. SPE is effective in humans with intestinal bowel disease (IBD) and chronic liver diseases. In addition, hypergammaglobulinemia is a marker of extraintestinal manifestations in human IBD patients, which often includes hepatic and pancreatic disease. Concurrent pancreatic and/or hepatic diseases in cats with chronic enteropathy (CE) have often been found, which may contribute to SPE alterations. The present study investigated the relationship between SPE and endoscopy, histopathology, and hepatic and pancreatic ultrasonographic findings in ninety-nine cats diagnosed with CE. No significant differences were observed between SPE and endoscopic and histopathological severity scores. Cats with concurrent pancreatic and/or hepatic ultrasonographic alterations showed lower albumin, lower α-globulin, and higher γ-globulin levels than cats diagnosed with enteropathy alone. This study suggests that hepatic and/or pancreatic alterations may influence SPE fractions in cats with CE, and should form the basis for further prospective studies on cats with diaditis and triaditis.Few studies have investigated total protein (TP) and serum protein electrophoresis (SPE) in cats with chronic enteropathy (CE). Cats diagnosed with CE were evaluated to investigate the relationships between TP, SPE and endoscopy, histopathology, and extraintestinal involvement. Medical records were searched for cats with a history of chronic gastrointestinal signs and a final diagnosis of CE. Information on signalment, TP, SPE, endoscopic score, histopathological diagnosis and score, and concurrent hepatic or pancreatic ultrasonographic alterations was collected. Relationships between protein profiles and other variables were investigated. Ninety-nine cats were included in the study, 63 diagnosed with various degrees of bowel inflammation and 36 with small-cell alimentary lymphoma. The most common TP alteration was hypoproteinemia (24%). No significant differences were observed between protein profiles and endoscopic and histopathological severity scores. Forty-five cats showing concurrent pancreatic and/or hepatic ultrasonographic alterations, had significantly lower albumin, lower α-globulin, and higher γ-globulin levels than cats not showing concurrent alterations. Disease severity scores did not seem to influence the protein profile in cats with CE. Extraintestinal involvement may be suspected in cats with lower albumin and α-globulins and higher γ-globulins.

A tiered approach to investigate the inhalation toxicity of cobalt substances. Tier 3: Inflammatory response following acute inhalation exposure correlates with lower tier data

In vitro studies have shown that cobalt substances predominantly induce pre-inflammatory biomarkers, resulting in a grouping of substances either predicted to cause inflammation following inhalation, or those with a different reactivity profile (poorly-reactive). There is a lack of data on whole-organ lung responses following inhalation of these substances, especially relating to the poorly-reactive group. It is of interest to generate tissue-specific histopathological correlation to better ascertain the predictive nature of the lower tier tests (i.e. tier 1 – bioelution, tiers 2a and b – in vitro markers and ToxTracker testing), in order to understand the type of effects caused by the poorly-reactive group and to gauge long-term effects. Eight cobalt substances were tested in vivo in a customized 4-h toxicity test; with animals sacrificed up to 14-days post-exposure. Histopathological severity scores were assigned based on inflammatory and pre-carcinogenic markers. A clear pattern emerged, with the reactive substances causing a persistent increase in one or more of the selected markers, and absence of these markers with poorly-reactive substances. Longer-term studies should be conducted to investigate the repeated dose effects of the poorly-reactive substances.

MO244CD47 BLOCKADE AMELIORATES AUTOIMMUNE VASCULITIS VIA THE EFFEROCYTOSIS OF NEUTROPHIL EXTRACELLULAR TRAPS*

Abstract Background and Aims Anti-neutrophil cytoplasmic antibody (ANCA)-associated vasculitis (AAV) is characterized by systemic necrotizing vasculitis in small vessels. The necrotic lesions consist of ANCA-mediated neutrophil extracellular traps (NETs) which represent a form of lytic cell death. The persistent NETs serve as autoantigens against ANCAs and cause organ damage in a vicious cycle. Considering dead cells are essentially cleared by phagocytic cells as a process of efferocytosis, why the NETs persist in tissue remains unclear. During efferocytosis, macrophages engulf apoptotic cells to prevent the leakage of intracellular components including toxic enzyme into the surrounding cells and these processes are regulated by the expression of CD47 as a “don’t eat me” signal. In this study, we hypothesized that ANCA-mediated NETs in AAV escape from efferocytosis via the up-regulation of CD47 and the persistent NETs amplify the disease. Method Human data: Human kidney biopsy specimens from patients with AAV and minor glomerular abnormality (MGA, as a case control) were subjected to immunohistochemistry (IHC) staining for CD47. In vitro: The expression of CD47 on neutrophils was evaluated by flow cytometry (FCM). Human neutrophils from healthy donor were treated with ANCA-IgGs from MPO-AAV patients or control IgGs. For the efferocytosis assay, macrophages were co-incubated with unstimulated, apoptotic, and ANCA-IgGs treated neutrophils in the presence of anti-CD47 monoclonal antibody (mAb) or a control antibody. The neutrophils were labeled with CFMDA cell tracker (fluorescent probe) and the efferocytosis was evaluated as neutrophil engulfed (CFMDA positive) macrophages using fluorescent microscopy. In vivo: Spontaneous crescentic glomerulonephritis-forming/Kinjoh (SCG/Kj) mice (8-week-old age) were treated with intraperitoneal injection of anti-CD47 mAb or a control antibody every 5 days for two weeks. The severity of glomerulonephritis was assessed by the levels of serum creatinine, haematuria, mRNA expression of pro-inflammatory genes, and histopathological findings. To address the immune response against the CD47 blockade, the titre of MPO-ANCA and the number of splenic cell subset was assessed by ELISA and FCM analysis, respectively. Results Human data: The IHC analysis of human renal specimens revealed that the positive area of CD47 of AAV was greater than that of MGA. In particular, the CD47-overexpressed cells were seen in glomeruli with necrotic crescent formation. In vitro: Mean fluorescence intensity (MFI) of CD47 in ANCA-IgGs treated neutrophils (NETs) was significantly higher than that in control IgGs treated neutrophils (ANCA-IgG; 442±21.4 a.u. vs control IgG; 402±10 a.u., p<0.05). In efferocytosis assay, apoptotic neutrophils were engulfed by macrophages (efferocytosis rate/ apoptotic neutrophil; 20.5±3.8%, live neutrophils; 0.9±0.5%). The efferocytosis rate of ANCA-induced NETs significantly decreased compared to apoptotic neutrophil, but anti-CD47 mAb improved the efferocytosis of ANCA-NETs (efferocytosis rate/ anti-CD47 mAb; 19.1±4.2%, control antibody; 7.7±2.2%, p<0.05). In vivo: the renal histopathological severity score, serum creatinine level of AAV mice treated with anti-CD47 mAb decreased compared to that of AAV mice treated with a control antibody (anti-CD47 mAb; 0.96±0.30 vs control antibody; 0.61±0.32 mg/dL). Although there was no significant difference in the number of splenic cells between anti-CD47 and control antibody treated mice, CD47 blockade therapy significantly reduced serum MPO-ANCA titre (28.5±10.4 vs 45.2±14.5 μg/mL) and renal mRNA expression (IFNα, IFNγ, MCP-1 and perforin) of AAV mice. Conclusion ANCA-mediated NETs might escape from efferocytosis through up-regulation of CD47 and provoke necrotizing vasculitis. CD47 blockade could be a potential novel therapeutic strategy for AAV.

Parametric Imaging of Contrast-Enhanced Ultrasound (CEUS) for the Evaluation of Acute Gastrointestinal Graft-Versus-Host Disease.

In recent years contrast-enhanced ultrasound (CEUS) has been an emerging diagnostic modality for the detection of acute gastrointestinal (GI) graft-versus-host disease (GvHD) in patients after allogeneic stem cell transplantation. However, broad clinical usage has been partially limited by its high dependence on the expertise of an experienced examiner. Thus, the aim of this study was to facilitate detection of acute GI GvHD by implementing false color-coded parametric imaging of CEUS. As such, two inexperienced examiners with basic knowledge in abdominal and vascular ultrasound analyzed parametric images obtained from patients with clinical suspicion for acute GvHD in a blinded fashion. As diagnostic gold standard, histopathological GvHD severity score on intestinal biopsies obtained from lower GI tract endoscopy was performed. The evaluation of parametric images by the two inexperienced ultrasound examiners in patients with histological confirmation of acute GI GvHD was successful in 17 out of 19 patients (89%) as opposed to analysis of combined B-mode ultrasound, strain elastography, and CEUS by an experienced examiner, which was successful in 18 out of 19 of the patients (95%). Therefore, CEUS with parametric imaging of the intestine was technically feasible and has the potential to become a valuable diagnostic tool for rapid and widely accessible detection of acute GvHD in clinical practice.

Nephroprotective Potential Effect of Azilsartan in Renal Ischemia Reperfusion Injury /role VEGF Pathway

The impairment of blood flow to the kidney causes renal injury occurs by ischemia/reperfusion in surgical procedures, in which the kidneys remainwithout blood supply for some time. This is observed during kidney transplantation, vascular surgery of aorta and renal arteries, and in partial nephrectomy. Inflammation is major contributor to the pathogenesis of renal ischemia/reperfusion in acute kidney injury with an endothelium loss, causing the vascular dysfunction and the angiogenesis deregulation. Azilsartan is an angiotensin II receptor antagonist used in the treatment of hypertension. It demonstrates organ protective effects in hypertension. In this study, we aim to assess the ability of azilsartanto exert potential protective effects on the kidney I/R and subsequent kidney dysfunctions in the rat through the modulation of pro‑inflammatory cytokinesand angiogenesis parameters (vascular endothelial growth factor receptor2 /vascular endothelial growth factor pathway). A total of 24 rats were randomly distributed into 4 groups (6 rats in each one) then they used in this experiment and the ischemia-reperfusion injury was induced in a rat model by bilateral renal artery clamping for 30 min and reperfusion for 2 h, and azilsartan (0.4 mg/kg) and vehicle was injected through intraperitoneally route before 30 min. prior of ischemia-reperfusion injury induction. Azilsartan administration could exert a protective role in the kidney against injury by significantly reducing the tissue levels of pro‑inflammatory cytokines (interleukin 1 beta, Monocyte chemoattractant protein-1and high mobility group box protein 1), lessen the serum levels of renal function parameters (blood urea nitrogen and serum creatinine), reduction of the histopathological severity score of renal damage and up regulation the angiogenesis markers (Vascular endothelial growth factor receptor 2, caveolin-1 and cluster of differentiation 34). All changes in the study parameters were significant (p value≤ 0.05). How to Cite this Article Pubmed Style Aseel Noaman Ali, Murooj L. Altimimi, Haider Mahdi Al-Ardi,Najah R Hadi. Potential Effect of Azilsartan in Renal Ischemia Reperfusion Injury /role VEGF Pathway. SRP. 2019; 10(2): 90-99. doi:10.5530/srp.2019.2.15 Web Style Aseel Noaman Ali, Murooj L. Altimimi, Haider Mahdi Al-Ardi,Najah R Hadi. Potential Effect of Azilsartan in Renal Ischemia Reperfusion Injury /role VEGF Pathway. http://www.sysrevpharm.org/?mno=302644928 [Access: March 31, 2021]. doi:10.5530/srp.2019.2.15 AMA (American Medical Association) Style Aseel Noaman Ali, Murooj L. Altimimi, Haider Mahdi Al-Ardi,Najah R Hadi. Potential Effect of Azilsartan in Renal Ischemia Reperfusion Injury /role VEGF Pathway. SRP. 2019; 10(2): 90-99. doi:10.5530/srp.2019.2.15 Vancouver/ICMJE Style Aseel Noaman Ali, Murooj L. Altimimi, Haider Mahdi Al-Ardi,Najah R Hadi. Potential Effect of Azilsartan in Renal Ischemia Reperfusion Injury /role VEGF Pathway. SRP. (2019), [cited March 31, 2021]; 10(2): 90-99. doi:10.5530/srp.2019.2.15 Harvard Style Aseel Noaman Ali, Murooj L. Altimimi, Haider Mahdi Al-Ardi,Najah R Hadi (2019) Potential Effect of Azilsartan in Renal Ischemia Reperfusion Injury /role VEGF Pathway. SRP, 10 (2), 90-99. doi:10.5530/srp.2019.2.15 Turabian Style Aseel Noaman Ali, Murooj L. Altimimi, Haider Mahdi Al-Ardi,Najah R Hadi. 2019. Potential Effect of Azilsartan in Renal Ischemia Reperfusion Injury /role VEGF Reviews in Pharmacy, 10 (2), 90-99. doi:10.5530/srp.2019.2.15 Chicago Style Aseel Noaman Ali, Murooj L. Altimimi, Haider Mahdi Al-Ardi,Najah R Hadi. Nephroprotective Potential Effect of Azilsartan in Renal Ischemia Reperfusion Injury /role VEGF Pathway. Systematic Reviews in Pharmacy 10 (2019), 90-99. doi:10.5530/srp.2019.2.15 MLA (The Modern Language Association) Style Aseel Noaman Ali, Murooj L. Altimimi, Haider Mahdi Al-Ardi,Najah R Hadi. Nephroprotective Potential Effect of Azilsartan in Renal Ischemia Reperfusion Injury /role VEGF Pathway. Systematic Reviews in Pharmacy 10.2 (2019), 90-99. Print. doi:10.5530/srp.2019.2.15 APA (American Psychological Association) Style Aseel Noaman Ali, Murooj L. Altimimi, Haider Mahdi Al-Ardi,Najah R Hadi (2019) Potential Effect of Azilsartan in Renal Ischemia Reperfusion Injury /role VEGF Reviews in Pharmacy, 10 (2), 90-99. doi:10.5530/srp.2019.2.15

A Multiplatform Approach for the Discovery of Novel Drug-Induced Kidney Injury Biomarkers.

Drug-induced kidney injury (DIKI) is a common toxicity observed in pharmaceutical development. We demonstrated the use of label-free liquid chromatography-mass spectrometry (LC-MS) and multiplex liquid chromatography-single reaction monitoring (LC-SRM) as practical extensions of standard immunoassay based safety biomarker assessments for identification of new toxicity marker candidates and for improved mechanistic understanding. Two different anticancer drugs, doxorubicin (DOX) and cisplatin (cis-diamminedichloridoplatinum, CDDP), were chosen as the toxicants due to their different modes of nephrotoxicity. Analyses of urine samples from toxicant treated and untreated rats were compared to identify biochemical analytes that changed in response to toxicant exposure. A discovery (label-free LC-MS) and targeted proteomics (multiplex LC-SRM) approach was used in combination with well established immunoassay experiments for the identification of a panel of urinary protein markers related to drug induced nephrotoxicity in rats. The initial generation of an expanded set of markers was accomplished using the label-free LC-MS discovery screen and ELISA based analysis of six nephrotoxicity biomarker proteins. Diagnostic performance of the expanded analyte set was statistically compared to conventional nephrotoxicity biomarkers. False discovery rate (FDR) analysis revealed 18 and 28 proteins from the CDDP and DOX groups, respectively, exhibiting significant differences between the vehicle and treated groups. Multiplex SRM assays were constructed to more precisely quantify candidate markers selected from the discovery screen and immunoassay experiments. To evaluate the sensitivity and specificity for each of the candidate biomarkers, histopathology severity scores were used as a benchmark for renal injury followed by receiver-operating characteristic (ROC) curve analysis on selected biomarkers. Further examination of the best performing analytes revealed relevant biological significance after consideration of anatomical localization and functional roles. In summary, the inclusion of mass spectrometry together with conventional ELISA based assays resulted in the identification of an expanded set of biomarkers with a realistic potential for providing additional beneficial information in mechanistic investigations of drug induced kidney injury and with similar responsiveness to conventionally applied indicators of renal injury.

Adiponectin exerts a potent anti-arthritic effect and insulin resistance in collagen-induced arthritic rats.

Previous research has shown that adiponectin (AD) induces severe insulin resistance (IR) and exhibits pro-inflammatory effect, so it could serve as a useful risk biomarker in rheumatoid arthritis (RA). The present study aims to evaluate the effect of AD on IR and anti-arthritis in collagen-induced arthritic (CIA) rats. After immunization with bovine type II collagen (CII), Wistar rats were administered with AD (60μg/kg/day) or saline into the ankle joint cavity of the left hind leg for 15days. The severity of arthritis was clinically and histologically assessed. Arthritis score was recorded every other day for each paw. Paw volume was measured on alternate days to monitor the progression of the disease in the arthritic control group. Tumor necrosis factor (TNF)-α, interleukin (IL)-1, AD, insulin and fasting glucose were measured in sera. Histopathology of joint synovial tissues was also examined. Treatment with AD resulted in significantly delayed onset of arthritis as well as decreased clinical arthritis and histopathological severity scores. AD reduced both serum fasting glucose, TNF-α, IL-1 and IR. Histological analysis confirmed treatment with AD suppressed joint synovial inflammation and immunohistochemical expression of TNF-α compared to the CIA group. Surprisingly, adiponectin levels measured by enzyme-linked immunosorbent assay in serum were significantly increased in CIA rats compared to the normal group. Adiponectin might display anti-inflammatory effects. These results suggest that AD may be a potential immunosuppressant for the treatment of RA linked to metabolic disease.

Micro-RNA expression in muscle and fiber morphometry in myotonic dystrophy type 1.

We aimed to explore the cellular action of micro-RNAs that are non-coding-RNAs modulating gene expression, whose expression is dysregulated in myotonic dystrophy (DM1). Basic procedure was to measure the levels of muscle-specific myo-miRNAs (miR-1, miR-133a/b, miR-206) in muscle of 12 DM1 patients. Muscle fiber morphometry and a new grading of histopathological severity score were used to compare specific myo-miRNA level and fiber atrophy. We found that the levels of miR-1 and miR-133a/b were significantly decreased, while miR-206 was significantly increased as compared to controls. The histopathological score did not significantly correlate with the levels of myo-miRNAs, even if the lowest levels of miRNA-1 and miRNA-133a/b, and the highest levels of miRNA-206 were observed in patients with either severe histopathological scores or long disease duration. The histopathological score was inversely correlated with disease duration. Nowadays that DM1 muscle biopsies are scanty, since patients are usually diagnosed by genetic analysis, our study offers a unique opportunity to present miRNA expression profiles in muscle and correlate them to muscle morphology in this rare multisystem disorder. Our molecular and morphologic data suggest a post-transcriptional regulatory action of myo-miRNA in DM1, highlighting their potential role as biomarkers of muscle plasticity.

Circulating microRNA 216 as a Marker for the Early Identification of Severe Acute Pancreatitis

BackgroundTo study the value of circulating microRNA 216 (miR-216) as a marker for the severity of acute pancreatitis (AP) in both murine models and patients. Materials and MethodsMice with AP were induced by intraperitoneal injection of 50μg/kg/hour cerulean either 7 times, sacrificed at 8, 9, 10, 11 or 12 hours after the first injection, or 12 times, sacrificed at 24 hours after the first injection. Plasma samples and data from patients with AP were obtained from a prospective cohort. Quantitative reverse transcription polymerase chain reaction was used to determine the miR-216a and miR-216b level. ResultsThe upregulation of miR-216a and miR-216b in the serum of mice was induced by cerulean injection in both the 7- and 12-injection groups (P < 0.05). The downregulation of miR-216a in pancreatic tissues of mice with AP was detected (P < 0.05), but no difference was observed in pancreatic miR-216b levels among any of the groups (all P > 0.05). The serum miR-216a level was positively correlated with pancreatic histopathology severity scores, and was negatively correlated with pancreatic miR-216a (r = −0.483, P = 0.009). The plasma miR-216a level was significantly upregulated in patients with severe AP (SAP) compared with patients with mild AP (MAP) or moderate severe AP (MSAP) (SAP versus MAP, P = 0.04; SAP versus MSAP, P = 0.00), but no difference was seen between patients with MAP and those with MSAP (P = 0.73). ConclusionsCirculating miR-216a might be a potential biomarker for the early identification of SAP.

Temporal kinetics of organ damage in copper toxicity: A histopathological correlation in rat model

Excess of copper is toxic to different organs. We aim to study the histopathological changes of liver, kidney, and brain following oral CuSO4 exposure for different duration and doses in rat model. Fifty-four males Wistar rats (205 ± 10 g) were included and divided into control (group-I) and experimental (group-II and III) arms. An oral dose of 100 and 200 mg/kgBWt/Day CuSO4 was given to group-II and III respectively and group-I received normal saline by gavage. Six rats from each group were sacrificed on days 30, 60 and 90 for biochemical and histopathological examinations. The histopathological changes were graded on 1–5 scores and correlated with respective laboratory parameters. The organ functions were worsened in experimental group with increasing dose and time. Histopathological study revealed edema, hemorrhage, necrosis and fibrosis/gliosis in experimental group. The worst histopathological severity score ranged from 4 to 5(median 5) in liver, 3–5(median 4) in kidney and 4–5(median 5) in brain. The edema and hemorrhage were more marked at 30 days and fibrosis/gliosis at 90 days. In conclusion, high-dose Cu toxicity results in structural damage to liver, kidney, and brain that correlates with organ dysfunction, Cu, GSH, TAC, and MDA concentrations. Liver damage is more severe and occurs earlier than other organs.

In response: challenges for statistical evaluation of ecotoxicological experiments--an industry perspective.

In response: challenges for statistical evaluation of ecotoxicological experiments--an industry perspective.

Selection of non-steroidal anti-inflammatory drug and treatment regimen for sulfur mustard-induced cutaneous lesions

The inflammatory process plays an important role in sulfur mustard (HD) injury and HD pathogenesis, suggesting that anti-inflammatory treatments applied as soon as possible following HD injury may reduce tissue damage and accelerate healing. This study used the HD dermal weanling swine model to investigate the efficacy of two non-steroidal anti-inflammatory drugs, capsaicin and diclofenac, when applied in combination with the steroid, clobetasol. The therapeutic regimen was also investigated with respect to initiation of treatment post-exposure, frequency and duration. Yorkshire-cross pigs were randomly assigned to experimental groups, corresponding to all combinations of treatment (capsaicin with clobetasol or diclofenac with clobetasol), onset time (1, 2 or 4 h post-exposure), treatment duration (1, 3 or 5 days) and frequency of applications (2, 3 or 4 per day). For each animal, two sites on the ventral abdomen were exposed to 400 μL of neat HD for 8 min to achieve superficial dermal (SD) lesions and two sites were exposed to 400 μL neat HD for 30 min to achieve deep dermal (DD) lesions. Each treatment regimen was tested against a SD and a DD injury. Untreated SD and DD lesion sites served as within-animal controls. Assessments, up to one week post-challenge, included digital photographs, clinical assessments (lesion size measurements and modified Draize scoring), transepidermal water loss (TEWL), reflectance colorimetry and histopathologic evaluations that included an estimate for depth of injury and wound healing parameters. Diclofenac plus clobetasol treatment resulted in significant reductions in lesion contracture and modified Draize scores, increased barrier function (decreased TEWL), and increased healing as determined by histopathology for both SD and DD injury when compared with untreated sites and sites treated with capsaicin plus clobetasol. An increased duration of treatment from 1 to 5 days was most commonly associated with decreased clinical assessment and histopathological severity scores. Therefore, a combination of diclofenac and clobetasol application, when administered for at least five days, shows promise in ameliorating HD-induced lesions.

Urinary and faecal N-methylhistamine concentrations do not serve as markers for mast cell activation or clinical disease activity in dogs with chronic enteropathies.

BackgroundThis study sought to correlate faecal and urinary N-methylhistamine (NMH) concentrations with resting versus degranulated duodenal mast cell numbers in dogs with chronic enteropathies (CE), and investigate correlations between intestinal mast cell activation and clinical severity of disease as assessed by canine chronic enteropathy clinical activity index (CCECAI), and between urinary and faecal NMH concentrations, mast cell numbers, and histopathological scores. Twenty-eight dogs with CE were included. Duodenal biopsies were stained with haematoxylin and eosin (H&E), toluidine blue, and by immunohistochemical labelling for tryptase. Duodenal biopsies were assigned a histopathological severity score, and duodenal mast cell numbers were counted in five high-power fields after metachromatic and immunohistochemical staining. Faecal and urinary NMH concentrations were measured by gas chromatography–mass spectrometry.ResultsThere was no correlation between the CCECAI and faecal or urinary NMH concentrations, mast cell numbers, or histopathological score – or between faecal or urinary NMH concentration and mast cell numbers. Post hoc analysis revealed a statistically significant difference in toluidine blue positive mast cells between two treatment groups (exclusion diet with/without metronidazole versus immunosuppression (IS)), with higher numbers among dogs not requiring IS.ConclusionFaecal and urinary NMH concentrations and duodenal mast cell numbers were not useful indicators of severity of disease as assessed by the CCECAI or histological evaluation. The number of duodenal mast cells was higher in dogs that did not need IS, i.e. in dogs responding to an exclusion diet (with/without metronidazole), than in dogs requiring IS. Further studies comparing the role of mast cells in dogs with different forms of CE are needed.

The critical threshold of Lawsonia intracellularis in pig faeces that causes reduced average daily weight gains in experimentally challenged pigs

Serology indicates that Lawsonia intracellularis infection is widespread in many countries, with most pigs seroconverting before 22 weeks of age. However, the majority of animals appear to be sub-clinically affected, demonstrated by the low reported prevalence of diarrhoea. Production losses caused by sub-clinical proliferative enteropathy (PE) are more difficult to diagnose, indicating the need for a quantitative L. intracellularis assay that correlates well with disease severity. In previous studies, increasing numbers of L. intracellularis in pig faeces, quantified with a real time polymerase chain reaction (qPCR), showed a strong negative correlation with average daily gain (ADG).In this study, the association between faecal L. intracellularis numbers and PE severity was examined in two L. intracellularis experimental challenge trials (n1=32 and n2=95). The number of L. intracellularis shed in individual faeces was determined by qPCR on days 0, 7, 14, 17 and 21 days post challenge, and average daily gain was recorded over the same period. The severity of histopathological lesions of PE was scored at 21 days post challenge. L. intracellularis numbers correlated well with histopathology severity and faecal consistency scores (r=0.72 and 0.68, respectively), and negatively with ADG (r=−0.44). Large reductions in ADG (131g/day) occurred when the number of L. intracellularis shed by experimentally challenged pigs increased from 107 to 108L. intracellularis, although smaller ADG reductions were also observed (15g/day) when the number of L. intracellularis increased from 106 to 107L. intracellularis.