Histopathological Severity Research Articles

Comparison of the current abattoir surveillance system for detection of paratuberculosis in Australian sheep with quantitative PCR tissue strategies using simulation modelling

Abattoir surveillance for Johne’s disease monitoring in Australia has provided valuable feedback to producers about their flock’s disease status since its commencement in 1999. The current surveillance system relies on the identification of gross lesions in sheep carcases at an abattoir, followed by sampling and histopathology testing. This manual inspection system has not been adapted to meet the changing disease situation, as infection prevalence levels have declined over time due to vaccination. This simulation study compares the current system with two alternative approaches utilising a validated quantitative (q)PCR method for the detection of Mycobacterium avium subsp. paratuberculosis in tissues, with random systematic sampling either alone or in conjunction with sampling of a single carcass presenting gross lesions. Consigned sheep were randomly simulated as either infected or uninfected according to defined prevalence levels of infection, with varying histopathological lesion severity and the presence or absence of gross lesions. These sheep were then allocated into multiple ‘lines’ (group of sheep slaughtered together) within each consignment, with each line subjected to testing with the three sampling strategies for the estimation of line and flock (consignment) sensitivity. The line sensitivity described the proportion of infected lines that tested positive, whereas the flock sensitivity was the proportion of consignments from the simulated infected flocks that had one or more lines test positive for paratuberculosis infection. The tissue qPCR strategy with gross lesion detection achieved marginally higher line sensitivity than the current abattoir surveillance strategy. The simulation of unvaccinated infected flocks with low to moderate prevalence levels demonstrated similar flock sensitivity for all three sampling models. However, the current strategy had very low line sensitivity for the simulated vaccinated infected flocks when the infection prevalence level was <2%. There were substantial differences in flock sensitivity between the two tissue qPCR approaches and the current abattoir surveillance strategy for vaccinated infected flocks, whereas, only marginal differences in flock sensitivity were evident between the two tissue qPCR models. Our results demonstrate that the current strategy is not effective at identifying infected animals at very low infection prevalence levels. The tissue qPCR approach investigated in this study is better as it removes the reliance on meat inspectors to identify gross lesions and can also assist in identifying flocks that have subclinical infected sheep not displaying gross lesions. Therefore, the sheep industry may benefit from incorporating tissue qPCR for Johne's disease surveillance, however the logistics and costs of conducting this type of testing would need to be considered prior to implementing any changes.

Chicken bone marrow mesenchymal stem cells improve lung and distal organ injury

Mesenchymal stem cells (MSCs) are associated with pulmonary protection and longevity. We separated chicken bone marrow-derived mesenchymal stem cells (BM-MSCs); investigated whether BM-MSCs can improve lipopolysaccharide (LPS)-induced lung and distal organ injury; and explored the underlying mechanisms. Ninety-six male ICR (6 weeks old) mice were randomly divided into three groups: Sham, LPS, and LPS + MSC groups. The mice were intratracheally injected with 5 mg/kg LPS to induce acute lung injury (ALI). The histopathological severity of injury to the lung, liver, kidney, heart, and aortic tissues was detected. Wet/dry ratio, protein concentrations in bronchoalveolar lavage fluid (BALF), BALF cell counts, inflammatory cytokine levels in serum, inflammatory cytokine gene expression, and oxidative stress-related indicators were detected. In addition, a survival analysis was performed in sixty male ICR mice (6 weeks old, 18–20 g). This study used chicken BM-MSCs, which are easier to obtain and more convenient than other animal or human MSCs, and have MSC-associated properties, such as a colony forming ability, multilineage differentiation potential, and certain phenotypes. BM-MSCs administration significantly improved the survival rate, systemic inflammation, and the histopathological severity of lung, liver, kidney, and aortic injury during ALI. BM-MSCs administration reduced the levels of inflammatory factors in BALF, the infiltration of neutrophils, and oxidative stress injury in lung tissue. In addition, BM-MSCs administration reduced TRL4 and Mdy88 mRNA expression during ALI. Chicken BM-MSCs serve as a potential alternative resource for stem cell therapy and exert a prominent effect on LPS-induced ALI and extrapulmonary injury, in part through TRL4/Mdy88 signaling and inhibition of neutrophil inflammation and oxidative stress injury.

Clinical signs, duodenal histopathological grades, and serum high-mobility group box 1 concentrations in dogs with inflammatory bowel disease.

BackgroundInflammatory bowel disease (IBD) commonly occurs in dogs, but there is lack of information about potential biomarkers of clinical and histopathologic severity.ObjectiveTo examine the role of serum C‐reactive protein (CRP) and high‐mobility group box 1 (HMGB1) concentrations in dogs with IBD.AnimalsSeventeen dogs with IBD and 25 healthy dogs.MethodsIn this prospective study, duodenal histopathologic severity was graded, and the clinical severity of IBD was assessed by the canine IBD assessment index (CIBDAI) score in dogs with IBD. Serum CRP and HMGB1 concentrations were compared between IBD and healthy dogs and analyzed according to histopathologic grade in dogs with IBD. The correlations between serum CRP and HMGB1 concentrations and the CIBDAI score were evaluated.ResultsDogs with IBD had higher serum CRP (median [range] = 20.39 [1.53‐67.69] μg/mL vs 2.31 [0.17‐11.49] μg/mL; P < .001) and HMGB1 concentrations (0.44 [0.07‐1.58] ng/mL vs 0.05 [0.01‐0.25] ng/mL; P < .001) than healthy dogs. The serum HMGB1 concentration was higher in IBD dogs with a moderate to severe histopathologic grade (0.51 [0.30‐1.58] ng/mL, P = .03) than in those with a mild histopathologic grade (0.17 [0.07‐0.75] ng/mL). Serum CRP concentrations and CIBDAI score were positively correlated in dogs with IBD (r s = .49, P = .05).Conclusions and Clinical ImportanceSerum HMGB1 could be a potential biomarker for diagnosing IBD and might be indicative of histopathologic severity in dogs, whereas serum CRP might be an indicator of clinical severity.

Punctate Pattern and Pemphigus: Is There Any Evidence of Punctate Pattern Among Iranian Patients?

To examine the prevalence of this novel pattern among Iranian patients with pemphigus and peruse the relationship between the presence of a punctate pattern with clinical severity of disease and histopathological findings. One hundred recently diagnosed patients with pemphigus were enrolled. DIF evaluation and routine light microscopy were performed on their biopsy specimens. Disease severity was determined using the Pemphigus Disease Area Index. Serum samples were collected to measure autoantibody titers using enzyme-linked immunosorbent assay. All the samples evaluated by DIF showed a continuous linear pattern of intercellular IgG deposition, whereas none of them had a punctate pattern. Despite a significant correlation between the Pemphigus Disease Area Index score and autoantibody values, no association between histopathological findings and disease severity has been found. We could not detect any punctate pattern among Iranian patients with pemphigus. The importance of this pattern in the diagnosis of pemphigus might be different among patients with different ethnic and genetic factors.

Liver fibrosis is associated with carotid atherosclerosis in patients with liver biopsy-proven nonalcoholic fatty liver disease

Nonalcoholic fatty liver disease (NAFLD) is related to subclinical atherosclerosis. However, whether the severity of the disease (or which histopathological component) is associated with subclinical atherosclerosis remains controversial. This study aimed to investigate the association between the histopathological severity of NAFLD and carotid intima-media thickness (CIMT) in Japanese patients with liver biopsy-proven NAFLD. Maximum-CIMT (max-CIMT) was measured as an index of carotid atherosclerosis in 195 biopsy-proven NAFLD patients. A significant association was observed between the severity of fibrosis (but not steatosis, inflammation, and ballooning) and max-CIMT. Older age, male gender, hypertension, and advanced fibrosis were independently linked to max-CIMT ≥ 1.2 mm. The prevalence of max-CIMT ≥ 1.2 mm was significantly higher in the advanced fibrosis group than in the non-advanced fibrosis group (75.4% versus 44.0%; p < 0.01). Non-invasive liver fibrosis markers and scoring systems, including fibrosis-4 index, NAFLD fibrosis score, hyaluronic acid, and Wisteria floribunda agglutinin positive Mac-2-binding protein, demonstrated that the diagnostic performance for max-CIMT ≥ 1.2 mm was similar to that of biopsy-based fibrosis staging. In conclusion, advanced fibrosis is significantly and independently associated with high-risk CIMT. Non-invasive fibrosis markers and scoring systems could help estimate the risk of atherosclerosis progression in patients with NAFLD.

The upregulation of oncogenic miRNAs in swabbed samples obtained from oral premalignant and malignant lesions.

Oncogenic miRNAs upregulated in OSCC play a range of versatile roles in oral carcinogenesis. Oral potentially malignant disorders (OPMDs) are the antecedent lesions to oral squamous carcinoma (OSCC) and they require a definitive diagnosis and early intervention. This study hypothesizes the presence of aberrant oncogenic miRNA expression in swabbed oral lesions. The expression of miR-21, miR-31, miR-134, miR-146a, and miR-211 in swabbed samples from 36 dysplastic or hyperplastic OPMDs and 10 OSCCs, relative to respective normal mucosa within the same patient, is analyzed with qRT-PCR to develop a diagnosis. Upregulation of all tested miRNAs in OPMD and OSCC samples comparing to controls is found to have occurred. Receiver operating characteristics curve analysis shows that miR-31 gives the best diagnostic accuracy of 0.91 when differentiating OPMD/OSCC from controls. An analysis of miR-134 and miR-211 expression allows the discrimination of the dysplastic state associated with OPMD, while the use of expression of the combined miRNAs further improves the analytical performances when identifying the dysplastic state. The concordant upregulation of miR-21, miR-31, and miR-146a is found to occur during an early stage of OSCC carcinogenesis. This study demonstrates the upregulation of multiple oncogenic miRNAs in swabbed OPMD and OSCC samples. miRNA expression in swabbed collectives enables the differentiation between normal mucosa and OPMD/OSCC, independent of their histopathological severity. This conventional and convenient sampling tool, when coupled with an assessment of miR-31 expression, would seem to be an adjuvant approach to the diagnosis of OPMD and OSCC.

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&amp;lt;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&amp;lt;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.

Upregulation of signal transducer and activator of transcription 3 in dogs with chronic inflammatory enteropathies.

BackgroundIn inflammatory bowel disease (IBD) in humans, phosphorylated signal transducer and activator of transcription 3 (pSTAT3) is upregulated in mucosal epithelial cells and correlates with clinical severity.Hypothesis/ObjectiveTo investigate the expression pattern of pSTAT3 in the mucosa of dogs with chronic inflammatory enteropathy (CIE) and explore correlations between its expression and clinical and histopathological severity scoring.AnimalsTwenty‐eight canine CIE patients grouped into food‐responsive enteropathy (FRE; 9), steroid‐responsive enteropathy (SRE; 10), and protein‐losing enteropathy (PLE; 9). Ten healthy beagle dogs served as controls (CO).MethodsRetrospective case control study. Immunohistochemistry was used to detect pSTAT3 in canine duodenal mucosa samples.ResultsCompared to CO, SRE (P < .001) and PLE (P < .001) dogs had significantly higher pSTAT3 expression in the villus epithelium. The SRE group had a significantly higher expression in the villus lamina propria (VLP) compared to controls (P = .009). In the crypt epithelium (CE), all CIE dogs had significantly higher pSTAT3 expression (FRE, P = .002; SRE, P = .003; PLE, P < .001) compared to CO. In the lamina propria crypt region (CLP), dogs with FRE (P = .04) and SRE (P = .03) had significantly upregulated pSTAT3 compared to controls. A positive correlation was found between canine chronic enteropathy clinical activity index (CCECAI) scoring and pSTAT3 expression for both epithelial (rho = .541; P < .001) and crypt regions (rho = .32; P = .02).Conclusions and Clinical ImportancepSTAT3 is upregulated in CIE in dogs, correlates with clinical severity, and may be helpful as a clinical marker in dogs with CIE.

The Serum Uric Acid Level Is Related to the More Severe Renal Histopathology of Female IgA Nephropathy Patients.

Hyperuricemia is a significant risk factor for cardiovascular morbidity and chronic kidney disease progression. IgA nephropathy (IgAN) is a well-known primary glomerular nephropathy. Hyperuricemia is associated with a poor prognosis in IgAN patients. We evaluated the association of hyperuricemia with the histopathological severity of IgAN in male and female patients; 658 patients diagnosed with IgAN via kidney biopsy were initially included. Baseline patient data were collected by eight university hospitals affiliated with the College of Medicine of the Catholic University of Korea. Pathological features were independently evaluated by eight expert pathologists working in the hospitals, and the consensus was reached. Of the initial 658 patients, 517 were finally included (253 males and 264 females). Hyperuricemia was defined as a serum uric acid (UA) level >7.0 mg/dL for males and >5.6 mg/dL for females; 108 (42.7%) males and 95 (35.9%) females exhibited hyperuricemia. Compared to the patients with normal UA levels, the global glomerulosclerosis, segmental sclerosis, mesangial matrix expansion (MME), endocapillary proliferation (ECP), interstitial fibrosis (IF), and tubular atrophy (TA) scores were higher in hyperuricemic males and females. In multivariable linear regression, the serum UA level correlated significantly with the MME, ECP, IF, and TA scores of female IgAN patients only.

Systemic delivery of microRNA for treatment of brain ischemia

Brain ischemia is the second leading cause of death and the third leading cause of disability in the world. Systemic delivery of microRNA, a class of molecules that regulate the expression of cellular proteins associated with angiogenesis, cell growth, proliferation and differentiation, holds great promise for the treatment of brain ischemia. However, their therapeutic efficacy has been hampered by poor delivery efficiency of microRNA. We report herein a platform technology based on microRNA nanocapsules, which enables their effective delivery to the disease sites in the brain. Exemplified by microRNA-21, intravenous injection of the nanocapsules into a rat model of cerebral ischemia could effectively ameliorate the infarct volume, neurological deficit and histopathological severity.

N-asetilsistein ve ozon gibi antioksidan ilaçların staz zonu üzerindeki etkinliği

Title: The effectiveness of antioxidant drugs such as N-acetylcysteine and Ozone on the stasis zone. Short title: Effectiveness of N-acetylcysteine and Ozone on the stasis zone. Purpose: Stasis zone, the zone around the coagulation zone, is important for the prevention necrosis in burns. Various drugs were studied in literature to prevent necrosis in stasis zone. The previous researches have shown the effectiveness of Nacetylcysteine treatment on both remote organs and local side in thermal injury. Demonstration of the effectiveness of N acetylcysteine and Ozone on stasis zone prevention in thermal injury was the goal of this study. Methods: For this purpose, 24 4-month-old Wistar albino type female rats weighing 200±20 g were used in the study. Compound burning model was made in these rats. Next, the compound was applied to the combustion model to compare the effectiveness of N-acetylcysteine and Ozone. To identfy histopathological destruction severity a scoring method was applied. Malondialdhyede (MDA), Glutathione Peroxidase (GPx), Superoksid dismutase (SOD), and catalase (CAT) levels were analyzed in serum samples of the rats to find out the alterations in the balance between oxidatitaion and antioxidation system. Results: While the highest MDA levels were observed in group I, the decrease was significant in group II and III (p=0.03 and p<0.02). SOD levels were significantly higher in group III compared to group II (p<0.05). CAT levels were higher in group II and III than group I (p<0.001, p<0.001). GPx levels were higher in group II (p<0.001), and group III (p<0.001). Histopathological scores were lower in group III than group I (p=0.001). Conclusion: This study showed that both Ozone and N-acetylcysteine are effective agents in rescuing the stasis site. It was also concluded that Ozone might be more beneficial than N-acetylcysteine.

Low-Risk Human Leukocyte Antigen Genes and Mild Villous Atrophy Typify Celiac Disease With Immunoglobulin A Deficiency.

We aimed to establish if in celiac disease (CD) with immunoglobulin A deficiency (IgAD) duodenal histopathology is influenced by human leukocyte antigen (HLA)-DQB1∗02 alleles dosage. Clinical differences between patients with CD and patients with CD and IgAD (CD-IgAD) were also evaluated. Five hundred and sixteen CD and 16 patients with CD-IgAD, enrolled over the time of 8 years, took part in this study. The severity of duodenal histopathology and frequency of CD at-risk HLA class II genes were compared in patients with CD versus patients with CD-IgAD. HLA class II genotypes were subdivided into two categories of genetic risk: high: HLA-DR3/DR7, -DR3/DR3, -DR4/DR4 -DR3/DR4 and low: HLA-DR5/DR7, -DR3/X, -DR4/X and X/X, where X means neither -DR3 nor -DR4. Then, they were compared with two types of duodenal histopathology: 0, 1, 2 and 3a of mild villous atrophy (MVA) and 3b and 3c of severe villous atrophy (SVA) according to the Marsh-Oberhuber classification. Clinical data concerning gender, number of esophagogastroduodenoscopies (EGDs) and association with other autoimmune diseases were obtained from medical records. In comparison with CD, CD-IgAD showed an increased frequency of MVA (P < 0.0001). Furthermore, CD-IgAD with MVA showed an increase of HLA low-risk genotypes (P = 0.036) and half HLA-DQ2 heterodimers (P = 0.0443). Interestingly, CD-IgAD demanded an increased number of EGDs to reach the diagnosis of CD (P = 0.0104) and autoimmune liver diseases were more frequent compared to CD (P = 0.0049). CD-IgAD is associated with MVA, low-risk HLA class II genes, an increased number of EGDs and autoimmune liver diseases.

Effect of Zinc Sulphate (ZnSO4) on the Gall Bladder of Clarias batrachus (Linn. 1758)

Heavy metal exposure to biota has become a severe thread to the environment, as its bioaccumulation, toxicity and persistence is happing at an alarming rate. Heavy metal presence in water bodies has been reported by various researchers. Henceforth their exposure to aquatic organisms is certain. An investigation on the possible toxicological impact of Zinc sulphate on fish Clarias batrachus is carried out in laboratory. Histology of gall bladder was used as a marker for evaluation of toxicity after sub lethal exposure. Fish were exposed to sub-lethal concentrations of 5mg/L, 10mg/L and 15mg/L for 20 days. Simultaneous control was also maintained. Principle histological alterations were deformities in the normal structures of columnar epithelial line, muscular layer, and mucosal folds. 15mg/L and 20mg/L exposure of zinc sulphate altered the sub mucosa besides leading the formation of bile crystals, an indication of water loss. The severity of histopathology was exposure dependent. Histopathological alteration in gall bladder can be regarded as sensitive biomarkers of Zinc sulphate toxicological manifestations hence can be utilized for ecotoxicological biomonitoring of aquatic bodies.

Association of MSY haplotype background with nonobstructive azoospermia is AZF-dependent: A case-control study.

Identifying causal genes of spermatogenic failure on the male-specific region of Y chromosome (MSY) has been a challenging process. Due to the nonrecombining nature of MSY, haplotype-based approaches have recently been shown to be promising in identifying associated MSY haplogroups. We conducted an MSY analysis of nonobstructive azoospermia (NOA) patients in a case-control setting (N=278 and 105 respectively) to identify modal haplogroups strongly associated with NOA. Patients with AZF deletions (AZF+) and no AZF deletions (AZF-) were compared with the control group. Given the larger sample set of AZF- NOA patients, we further investigated the association based on histopathological severity, namely Sertoli cell-only syndrome and maturation arrest subtypes. We observed no significant enrichment of MSY haplogroups in AZF- azoospermic patients (or its subtypes). However, we observed a strongly significant association between haplogroup J2a* and AZF+patients (FDR-corrected p=.0056; OR=7.02, 95%CI 1.89 to 39.20), a haplogroup which also showed significant enrichment for AZFa/b deletions (p=4x10-4 ). We conclude that unlike AZF+patients, AZF- NOA are less likely to have an MSY causative factor with large effect size, thus indicating that the aetiology of AZF- NOA, and to some extent AZFc NOA, is more likely to be based on non-MSY factors.

Colonoscopic evaluation of diarrhea/colitis occurring as an immune-related adverse event.

Diarrhea is often observed as an immune-related adverse event. In this study, we conducted a retrospective review of the severity of diarrhea, its treatment and the endoscopic findings in patients developing diarrhea as an immune-related adverse event. From August 2015 to June 2019, a total of 369 patients received treatment with immune checkpoint inhibitors at our hospital. For this study, development of grade 2 or more diarrhea in these patients was defined as an immune-related adverse event. We analyzed the histopathological severity of the bowel lesions according to the Nancy histological index for ulcerative colitis. Of the 369 patients, 27 (7.3%) developed diarrhea as an immune-related adverse event. Of these 27 patients, 18 received steroid treatment. Colonoscopy was performed in 17 patients and culture of the feces in 18. The tests revealed evidence of bacterial colitis (Aeromonas hydrophila) in two patients. The Nancy histological index was 4, 3, 2, 1 and 0 in two, three, two, two and seven patients, respectively. No findings on colonoscopy were observed in 7 of the 17 patients (41%) who underwent colonoscopy, and most of these patients recovered without steroid treatment. Patients with lower values of the Nancy histological index tended to show better responses to steroid treatment. To avoid unnecessary steroid administration, colonoscopic evaluation is essential in patients receiving treatment with immune checkpoint inhibitors who present with diarrhea as an immune-related adverse event. In addition, the endoscopic findings could be useful to predict the response to steroid treatment.

Human Umbilical Cord Blood Derived-Mesenchymal Stem Cells Alleviate Dextran Sulfate Sodium-Induced Colitis by Increasing Regulatory T Cells in Mice.

Inflammatory bowel disease (IBD), which main clinical manifestations include abdominal pain and diarrhea occurring repeatedly, is a kind of autoimmune disease. It has been reported in preceding studies that mesenchymal stem cells (MSCs) can reduce inflammation by regulating the function of immune cells. But studies about the interaction between MSCs and adaptive immune cells, especially in IBD models, are insufficient. Therefore, the objective of this research was to estimate the therapeutic effects of MSCs from human umbilical cord blood (hUCB-MSCs) in an IBD model of rodent and to clarify the therapeutic mechanisms of hUCB-MSCs. Dextran sulfate sodium (DSS) was used to induce colitis in rodent. Mice with colitis were treated with intraperitoneal infusions of hUCB-MSCs and evaluated for mortality and diverse disease symptoms containing weight reduction, diarrhea, and bloody stools. The levels of histopathologic severity and generation of regulatory T cells (Treg) were also determined. Treatment with hUCB-MSCs ameliorated the clinical and histopathologic severity of acute and chronic colitis in mice. Furthermore, T cell infiltration into the inflamed colon was significantly decreased (p = 0.0175), and Foxp3+ cells were substantially higher in the hUCB-MSC group than that of the DSS group. Our results suggest that hUCB-MSCs are able to alleviate inflammation via adding Foxp3+ Tregs in an IBD model of mouse. As a result, these findings suggest the opportunity of hUCB-MSC being applied to patients with IBD.

Age-Related Changes in the Canine Aorta.

Degenerative changes in the aorta are commonly observed in both dogs and humans, and those changes that occur with age morphologically overlap with those observed in genetic or degenerative diseases. Therefore, recognition of age-related aortic changes is important for diagnosticians, as such histologic findings should be distinguished from lesions of specific diseases. The aortas from 37 dogs without clinical cardiovascular disease ranging in age from 2 months to 15 years were divided into 3 cohorts to assess age-relatedness, and evaluated histologically using standardized nomenclature and diagnostic criteria adapted and modified from the human literature. We found that the histopathologic severity scores for intimal thickening, translamellar medial fibrosis, loss of smooth muscle cell nuclei, and medial microcalcification were higher in older dogs, whereas the scores for both intralamellar and translamellar mucoid extracellular matrix accumulation ("cystic medial necrosis") were not different among age groups. Dogs with translamellar medial fibrosis and aortic medial microcalcification were significantly older compared with dogs without these findings, while the presence of aortic medial chondro-osseous metaplasia was not related to age. Taken together, we demonstrate a range of age-related aortic histologic changes in dogs without clinical cardiovascular disease and suggest that integration of signalment and clinical data can aid in the differentiation of such findings from non-age-related disease processes.

Identification of endoscopic factors that predict poor responses to steroids in patients with gastrointestinal acute graft-versus-host disease

Gastrointestinal acute graft-versus-host disease (aGVHD) is a life-threatening complication that requires urgent and appropriate treatment. An endoscopic examination is considered the gold-standard for the diagnosis of gastrointestinal aGVHD. However, the prognostic value of endoscopy remains controversial. This study aimed to investigate the usefulness of pre-treatment macroscopic and histopathologic findings of upper and lower endoscopy with respect to predicting steroid-resistant gastrointestinal aGVHD. This retrospective study included 44 patients with gastrointestinal aGVHD who underwent endoscopy at the time of diagnosis and received systemic steroid treatment at our hospital. We graded the macroscopic and histopathologic findings using a previously validated 4-point scale. Univariate analyses of endoscopic grading revealed that a higher macroscopic grade in the ileum and higher histopathologic grades in the ileum and colon predicted a poor response to systemic steroids. In a multivariate analysis, macroscopic and histopathologic severity in the ileum were identified as significant prognostic factors that indicated resistance to steroid therapy. The presence of granulation tissue was also a strong independent predictor of resistance to steroid therapy. These findings suggest that both macroscopic and histopathologic findings in the ileum may be useful predictors of steroid-refractory gastrointestinal aGVHD and can indicate an immediate need to develop a second-line strategy.

Role of free radical scavenging activity of vasoactive intestinal peptide in the attenuation of mitochondrial dysfunction to ameliorate dextran sulphate sodium‐induced colitis in mice: Implications in ulcerative colitis

To evaluate the efficacy of vasoactive intestinal peptide (VIP) in treating ulcerative colitis (UC), targeting colonic mitochondrial dysfunction by virtue of its free radical scavenging properties for maintenance of colon mucosal integrity. A murine model was administered with dextran sodium sulfate (DSS) to induce colitis in C57BL/6J mice at 3.5%/g bodyweight for 3 cycles of 5 days each, followed by an intraperitoneal dose of VIP at 0.5 nmol/L per mouse per day for 10 days. The post-treatment mice were sacrificed and their colon samples were utilized for further analysis. To substantiate the in vivo findings and identify the reactive species involved in progression of UC, Caco-2 cells were subjected to DSS (5%) for 24 hours at 37 °C with or without VIP (10 nmol/L) in the presence or absence of specific free radical scavengers and antioxidants. Treatment with VIP reduced histopathological severity of colitis and cell death markers in murine model, leading to partial recovery of inhibited mitochondrial respiratory complexes, altered mitochondrial membrane potential and lowered adenosine triphosphate generation. Interestingly, in vitro treatment with VIP restored mitochondrial functions and its efficacy was equal to super oxide dismutase and dimethyl sulfoxide, indicating involvement of superoxide free radical (O2 •-) and hydroxyl radical (•OH) in progression of UC. However, catalase, Nω-nitro-l-arginine methyl ester and mercaptoethylguanidine were ineffective, indicating non-involvement of hydrogen peroxide, nitric oxide and ONOO- in UC. By virtue of its free radical scavenging properties VIP can act as a potent anti-colitogenic agent, reversing colonic mitochondrial dysfunction for treating UC.