Direct Immunofluorescence Staining Research Articles

Navigating Blistering Pathways: A Case Study of Dyshidrosiform Bullous Pemphigoid

Dyshidrosiform Bullous Pemphigoid (DBP) is a rare type of autoimmune blistering disease, mainly affecting elderly individuals. Causing pruritic blisters on the palms and soles, it can be challenging to differentiate from other conditions such as pompholyx. A case study of an 89-year-old patient with DBP is presented, emphasizing the importance of accurate diagnosis through techniques like direct immunofluorescence (DIF) staining. Treatment typically involves corticosteroids, monoclonal antibodies, dapsone, or immunosuppressants. Improved awareness and understanding of DBP are crucial for improved patient outcomes.

High serum total IgE levels correlate with urticarial lesions and IgE deposition in perilesional skin of bullous pemphigoid patients: An observational study.

Background In the pathophysiology of bullous pemphigoid, besides IgG, there has been evidence that supports the role of IgE antibodies. However, there have been no studies to evaluate total serum IgE levels or detect IgE deposits in the skin of Vietnamese patients. Aim To analyse the association between IgE levels in the serum and disease severity as well as eosinophils and IgE basement membrane zone (BMZ) deposition in Vietnamese bullous bullous pemphigoid patients. Methods A single-centre observational research on 35 newly diagnosed and untreated bullous bullous pemphigoid patients. Total serum IgE levels were analysed using enzyme-linked immunosorbent assay (ELISA). For controls, we collected sera of 30 pemphigus patients and 30 elderly patients with pruritus. Perilesional skin biopsies underwent direct immunofluorescence (DIF) staining, with biopsies of pemphigus patients as controls. Results Elevated total serum IgE was observed in 60% of bullous pemphigoid patients, the percentage in the pemphigus group and pruritus group was 20% and 40%, respectively. The mean total serum IgE level among the bullous pemphigoid group was higher than that of the pemphigus group (123.3 ± 102.4 IU/mL vs. 64.3 ± 45.1 IU/mL, p = 0.010). Total serum IgE levels of bullous pemphigoid patients correlated with higher eosinophil counts (r = 0.61; p = 0.018) and urticaria/erythema (U/E) Bullous Pemphigoid Disease Area Index (BPDAI) score (r = 0.50; p = 0.035). Among 35 bullous pemphigoid patients, 5 patients showed positive IgE DIF staining, accounting for 14.3%. Higher serum IgE levels correlated with the deposition of IgE in patients' perilesional skin (p = 0.037). Limitations Due to the rarity of bullous pemphigoid, the effect of the COVID-19 pandemic, and self-treatment issues in Vietnam, we could not recruit a larger number of participants. Conclusions Total serum IgE values correlated with urticarial lesions and IgE deposition in perilesional skin of Vietnamese bullous pemphigoid patients. IgE autoantibodies present in the skin of bullous pemphigoid patients support the role of IgE in bullous pemphigoid pathogenesis.

Direct immunofluorescence (DIF) versus immunohistochemical (IHC) staining of complements and immunoglobulins (Ig) in pemphigus group.

Pemphigus is a group of bullous disorders of the skin characterized by the formation of autoantibodies present in the intercellular junction of the epidermis. Diagnosis is made by clinical, histopathological examination, and DIF. As DIF needs frozen sections, fluorescent tagged antibodies, UV light microscope for examination, and trained personnel, its non-availability makes a definitive diagnosis challenging. To evaluate the utility of IHC staining of complements and Ig in cases of Pemphigus. Twenty-six diagnosed cases of Pemphigus were stained by Peroxidase immunohistochemical method using monoclonal antibody to IgG, IgA, IgM, IgG4, C3, C4 d with DAB as chromogen. Pemphigus cases include twenty of pemphigus vulgaris (PV), four cases of pemphigus foliaceous (PF), and two of pemphigus vegetans (Pveg). Positivity was defined as the deposition of Ig and complements as distinct, continuous brown staining of keratinocytes at intercellular junctions. On IHC total of 20 PV 17 showed positivity (85%) for IgG, 11 (55%) C4d, 19 (95%) C3d, and 16 (80%) IgG4 deposits at the intercellular junction of the epidermis. All cases of PF showed a deposit of IgG, with three (75%) cases for IgG4, C3d, and C4d. Both cases of Pveg showed positivity for IgG and C4d while one case was negative for IgG4 and C3d. The overall IgG, C3, IgG4, and C4d expression for pemphigus was seen in 88%, 88%, 76.9%, and 61.5% of cases. The relation between these markers, combination of IgG and C3, was best related to each other ( P value = 0.80). The sensitivities for IgG, IgG4, and C3 were 77.8%%, 73%, and 73% resp. We conclude that IHC is a useful tool in the diagnosis of PV with the highest sensitivity of IgG and C3d. The combination of IgG and C3d could replace the DIF in almost all of our cases, so IHC on FFPE sections be used as an alternative method to DIF.

Sinomenine Hydrochloride Protects IgA Nephropathy Through Regulating Cell Growth and Apoptosis of T and B Lymphocytes.

Sinomenine hydrochloride (SH) is used to treat chronic inflammatory diseases such as rheumatoid arthritis and may also be efficacious against Immunoglobulin A nephropathy (IgAN). However, no trial has investigated the molecular mechanism of SH on IgAN. Therefore, this study aims to investigate the effect and mechanism of SH on IgAN. The pathological changes and IgA and C3 depositions in the kidney of an IgAN rat model were detected by periodic acid-Schiff (PAS) and direct immunofluorescence staining. After extracting T and B cells using immunomagnetic beads, we assessed their purity, cell cycle phase, and apoptosis stage through flow cytometry. Furthermore, we quantified cell cycle-related and apoptosis-associated proteins by Western blotting. SH reduced IgA and C3 depositions in stage 4 IgAN, thereby decreasing inflammatory cellular infiltration and mesangial injury in an IgAN model induced using heteroproteins. Furthermore, SH arrested the cell cycle of lymphocytes T and B from the spleen of IgAN rats. Regarding the mechanism, our results demonstrated that SH regulated the Cyclin D1 and Cyclin E1 protein levels for arresting the cell cycle and it also regulated Bax and Bcl-2 protein levels, thus increasing Cleaved caspase-3 protein levels in Jurkat T and Ramos B cells. SH exerts a dual regulation on the cell cycle and apoptosis of T and B cells by controlling cell cycle-related and apoptosis-associated proteins; it also reduces inflammatory cellular infiltration and mesangial proliferation. These are the major mechanisms of SH in IgAN.

Detection of zoonotic enteropathogens in captive large felids in Italy.

Within the One Health paradigm, infectious disease surveillance have been developed for domestic and wild animals, leaving the role of captive non-domestic populations, especially felids in zoos and circuses, less explored. This study addresses the proximity of these captive animals to urban areas, necessitating focused monitoring for potential zoonotic enteropathogens. The present work aimed to investigate the presence of such zoonotic enteropathogens in faecal samples from captive large felid populations. A total of 108 faecal samples were collected in three circuses, five zoos and one rescue centre across Italy. Salmonella spp. isolation, serotyping and antimicrobial susceptibility testing were conducted on all samples. Additionally, 60 samples were also examined for gastrointestinal parasites using standard coprological techniques. Giardia spp. detection employed direct immunofluorescent staining and specific PCR, while Toxoplasma gondii was detected using PCR targeting B1 gene. A total of 51 Salmonella enterica subsp. enterica were isolated, with predominant serovariants including Infantis (43.1%), Coeln (11.8%) and Newport (11.8%). The captive felids likely act as asymptomatic carriers of foodborne Salmonella, with notable resistance ampicillin and trimethoprim-sulfamethoxazole, no resistance to enrofloxacin was noted. Microscopic analysis revealed Toxascaris leonina eggs in 11 faecal samples (18.3%) and Giardia duodenalis cysts in one animal (1.7%). Captive animals in public settings may act as sources of Salmonella infection and enteroparasitosis for both occupational and general exposure. The study emphasizes the role of captive animals in antimicrobial resistance dynamics, highlighting the need for routine pathogen screening in the management practices of zoological structures.

Giardiasis in the Southwest Region of the United States: Causes and Preventive Measures

Giardia is a protozoan parasite of the small intestine, and a leading cause of diarrheal diseases worldwide in a variety of animals, as well as humans. Despite the decrease, Giardiasis remains the most commonly reported intestinal parasitic infection principally in the Southwestern region of the United States. The parasite is typically transmitted by fecal-oral route and may be transmitted by direct intake or indirectly from contaminated food or water. Symptomatic Giardiasis is often treated to reduce the duration of symptoms, to prevent future complications, and to minimize transmission of the parasite to other hosts. The clinical image of Giardiasis is mixed, with high variability in severity of clinical disease which can become chronic or be followed by post-infectious sequelae. At present, treatment options include nitroimidazoles derivatives; especially metronidazole, which has been the mainstay of treatment for decades and is still widely used. The Navajo Nation located in the Southwestern region of the United States has suffered from the scarcity of water expediting Navajo populations to use water resources such as underground water, river streams, lakes and surface water as their main source of accessibility. The lack of adequate domestic and municipal water has led several Navajo families to become ill with waterborne infectious diseases like Giardiasis. More recently, molecular methods have become amateur in diagnosis for testing patient samples as well as water samples for Giardiasis. Through direct immunofluorescent staining and other methods that detect intact organisms have improved the detection of Giardia trophozoites or cysts. An understanding of clinical and molecular variability of organisms is important for effective treatment and the response of remedies that are commonly used. The molecular classification of bacteria and viruses isolated from Indigenous patients might have their own specification that can be correlated with isolates from water samples collected from the same localities. This correlation could help us in source tracking and finding solutions for the communities who are already at risk due to the lack of infrastructure and resources. In this review of literature, the causes and preventive measures of Giardiasis are covered to potentially help understand the importance behind this parasitic disease, more specifically on the Navajo Nation of the Southwest of the US.

Histologic Features of Syphilitic Gastritis: A Rare but Resurging Imitator of Common Diseases.

The range of histopathologic features of gastric syphilis is not well described. Here we describe the clinicopathologic findings of eight patients with syphilitic gastritis. A search of our Pathology Data System (2003-2022) and multiple other institutions identified eight patients with syphilitic gastritis. Clinical information, pathology reports, and available slides were reviewed. Lesions predominated in middle-aged adults (mean age, 47.2 years; range, 23-61 years) with a propensity for men (n = 7). Three patients had a documented history of human immunodeficiency virus. Clinical presentations included weight loss, abdominal pain, hematochezia, fever, dyspepsia, nausea and vomiting, hematemesis, anemia, and early satiety. Endoscopic findings included ulcerations, erosions, abnormal mucosa, and nodularity. All specimens shared an active chronic gastritis pattern with intense lymphohistiocytic infiltrates, variable plasma cells, and gland loss. Prominent lymphoid aggregates were seen in four specimens. The diagnosis was confirmed either by immunostain for Treponema pallidum (n = 7) or by direct immunofluorescence staining and real-time polymerase chain reaction (n = 1). All patients with available follow-up data showed resolution of symptoms after antibiotic therapy (n = 4). Recognition of the histologic pattern of syphilitic gastritis facilitates timely treatment, prevents further transmission, and avoids unnecessarily aggressive treatment.

Bioactive compounds anthocyanins as a factor in the nutritional recovery of the body's adaptive potential after intense physical activity in the experiment: assessment of immunological and hematological indicators of adaptation

Restoring the adaptive potential of an athlete is of paramount importance not only for the implementation of his training and competitive activities, but also for maintaining health. One of the leading place in complex recovery programs in sports is given to full-fledged optimal nutrition, which provides for meeting the body's requirements not only in energy, macro- and micronutrients, but also in minor bioactive compounds. The use of anthocyanin-containing products is a promising strategy for the normalization of metabolic and immune disorders that develop as a result of intense physical and neuro-emotional stress not only in athletes, but also in other groups of people exposed to these factors, including military personnel undergoing training in conditions close to combat. This determines the relevance of this study. The aim of the research was to study the effect of an anthocyanin-enriched diet on hematological profile and cellular immunity in rats after intense physical activity. Material and methods. The experiment was carried out for 4 weeks on 4 groups of male Wistar rats with an initial body weight of ~300 g. The motor activity of the animals of the 1st (control) and 2nd groups was limited by the standard keeping animals in the vivarium, while physically active rats of the 3rd and 4th groups received additional physical activity - training on a treadmill. Before the end of the experiment, the animals of 3rd and 4th groups were given debilitating physical activity on a treadmill (until the rats refused to continue the exercise). Rats of all 4 groups received a standard semi-synthetic diet, water ad libitum. Animals in 2nd and 4th groups were additionally fed blueberry and blackcurrant extract (30% anthocyanins) as part of the diet at a daily dose of 15 mg anthocyanins/kg body weight. Hematological parameters were determined on a Coulter ACT TM 5 diff OV hematological analyzer. Expression of CD45R, CD3, CD4, CD8a, CD161 receptors on rat peripheral blood lymphocytes was determined by direct immunofluorescent staining of whole blood cells using a panel of monoclonal antibodies conjugated with fluorescent dyes: APC, FITC, PE. The measurements were carried out on an FC-500 flow cytometer. Results. Intense physical activity in rats of the 3rd group did not lead to a significant change in erythrocyte parameters compared with the control group. Enrichment of the diet with blueberry and black currant extract (the 2nd and the 4th groups) provided a significant (p<0.05) increase in blood content of hemoglobin (Hb) (150.7±0.9 and 154.4±2.0 vs 145.4±0.9 g/l in control), hematocrit (44.95±0.21 and 46.18±0.64 vs 43.78±0.32%) and the average content of Hb in erythrocytes (18.00±0.20 and 18.03±0.24 vs 17.35±0.24 pg). The absolute content of leukocytes and other cellular elements of the leukocyte formula, as well as leukocyte indices in rats of the experimental groups didn't significantly differ from those of the control rats, which confirms the absence of an inflammatory process. Intense physical activity and anthocyanin enrichment of the diet didn't have a significant effect on rat platelet parameters. Enrichment of the diet of rats of the 4th group with blueberry and black currant extract led to the activation of cellular immunity, as evidenced by a significant (p<0.01) increase in the percentage (from the total content of T-lymphocytes) of T-helpers (70.13 ±1.34 vs 63.75±0.99%) and a decrease in the relative content of cytotoxic T-lymphocytes (28.65±1.38 vs 34.71±0.95%) in comparison with those in rats of the 3rd group and at the level of the trend (р<0.1) - from the 1st group indexes (66.87±1.20 and 31.87±1.26%, accordingly). Intense physical activity led to a decrease in immunoregulatory index in rats of the 3rd group (1.86±0.07) compared with the control (2.13±0.12) (p<0.1), and in animals of the 4th group this indicator was significantly higher (2.50±0.14, p<0.05). In animals of the 3rd group a statistically significant (p<0.05) decrease in the relative content of NK cells in peripheral blood was found compared to the control. Enrichment of the diet of physically active rats with blueberry and black currant extract led to a significant (p<0.05) increase in the percentage of NK cells compared to this indicator in rats of the 3rd group (4.87±0.75 vs 2.08±0.18%) and had no significant difference with the indicator in rats of the control group (4.32±0.98%). Conclusion. The enrichment of the rats' diet with blueberry and blackcurrant extract containing a daily dose of 15 mg of anthocyanins per kg of body weight provides an increase in blood Hb content, hematocrit and the average content Hb in erythrocytes. It has been established that intense physical activity induces the cellular immunity suppression. The activating effect of anthocyanins on adaptive cellular immunity and NK cells, which are lymphocytes of innate immunity, was revealed. The data obtained indicate the effectiveness of the use of bioactive compounds (anthocyanins) to increase the adaptive potential of the organism.

First report on the isolation of Chlamydia abortus from female dromedary camels with ovarian hydrobursitis

This study sought to isolate Chlamydia abortus (C. abortus) from camels with ovarian hydrobursitis (OVHB). To accomplish this goal, bursal tissue (n = 5) and bursal fluid (n = 6) samples were collected from 11 female dromedary camels with unilateral OVHB. A quantitative polymerase chain reaction (qPCR) was used for the preliminary detection of C. abortus in the infected samples. For the purpose of isolation, the prepared samples were inoculated into embryonated chicken eggs. Giemsa, Gimenez, and direct immunofluorescence (DIF) staining were used to detect any chlamydial inclusions in the infected yolk sacs. A second qPCR was then performed on the infected yolk sacs. The C. abortus gene was detected in 83.8% of the infected bursal tissue and bursal fluid samples. All the yolk sac smears treated with Giemsa, Gimenez, and DIF staining revealed intracytoplasmic inclusion bodies. Moreover, hemorrhagic patches, massive congestion, macerated yolk sacs, and dwarfism were observed in the infected chicken embryos. The C. abortus gene was also found in 63.6% of the infected yolk sacs. In conclusion, this is the first report of C. abortus isolation from female dromedary camels with OVHB, which represents a key step toward developing a practical vaccine and avoiding fertility problems in female camels.

Progressive ascending telangiectasias

Progressive ascending telangiectasias

Leucocytoclastic Vasculitis Following Coronavirus Vaccine: A Case Report

Background: The coronavirus disease 2019 pandemic is an unprecedented situation. Mass vaccination is a considered a promising solution to combat this global health crisis. Different vaccine have used to control transmission of the disease. Some complications of this vaccine were reported including rare cases of vasculitis. We report a case of leucocytoclastic vasculitis following BNT162b2 corona virus vaccine. Case Presentation: A 42 year old man with no medical history or drug intake presented 3 days after the third dose of BNT162b2 vaccine an infiltrated purpura at the lower extremities, fever, asthenia and myalgia. At physical examination, multiple palpable indurated purpuric papules were present only on the lower extremities. Laboratory tests were normal. The skin biopsy revealed leucocytoclastic vasculitis. The direct immunofluorescence staining detected only fibrinogen deposition. Conclusion: Leucocytoclastic vasculitis is an exceptional vaccine side effect. The resemblance of the vaccine-induced spike proteins with human components is highly likely to produce pathological autoantibodies and vaccine-induced autoimmunity through molecular mimicry.

Fc-Mediated Complement Activation Is Associated with Macrophage Trafficking and Induction of Neutrophil Extracellular Traps in Transfusion-Related Acute Lung Injury

Transfusion-related acute lung injury (TRALI) is a leading cause of blood transfusion-related fatalities without available therapies. The pulmonary endothelium is damaged in TRALI, through incompletely understood pathogenic mechanisms, resulting in pulmonary edema. Generally, anti-leukocyte antibodies or biological response modifiers in the transfusion product, in combination with predisposing risk factors in the transfused recipient (e.g. inflammation), are responsible for initiation of TRALI. Remarkably, not all anti-leukocyte antibodies cause TRALI. In a previous in vitro study, we identified increased Fc-mediated complement activation to be a key feature of murine TRALI-inducing antibody 34-1-2S (anti-H-2K d) compared to TRALI-resistant antibody SF1.1.10 (anti-H-2K d) (Zeeuw van der Laan et al, Blood Adv 2020).In the current study, we further explored antibody-mediated TRALI mechanisms in vivo using our previously established TRALI mouse model in which mice are pre-depleted of protective CD4+ T cells and primed with LPS, followed by infusion of antibody 34-1-2S (Kapur et al, Blood 2017, Blood Adv 2018, Blood 2019). A key read-out for TRALI was the lung wet/dry weight ratio (Lung W/D, measure for pulmonary edema). We found that in vitro antibody-mediated complement activation was associated with in vivo antibody-mediated TRALI. 34-1-2S caused severe TRALI (complement activation: +++, Lung W/D: 7.4 ± 0.21), while SF1.1.10 did not cause TRALI (complement activation: +, Lung W/D: 4.68 ± 0.16). Furthermore, Fc-deglycosylated 34-1-2S still caused significant TRALI (complement activation: ++, in vivo Lung W/D: 5.16 ± 0.52). TRALI development was fully Fc-dependent as 34-1-2S-Fab did not cause any TRALI (no complement activation, Lung W/D: 4.35 ± 0.18). Importantly, we found significantly increased levels of complement C1q-C4 complexes in plasma samples of TRALI patients (n=46) compared to healthy controls (n=25): 6.49 ± 5.27 vs 3.84 ± 2.27 AU/ml, respectively, P: 0.0005,***. Similarly, C5a levels were significantly elevated in plasma samples of TRALI patients (n=53) compared to healthy controls (n=30): 2.52 ± 2.17 vs 1.37 ± 1.08 ng/ml, respectively, P: 0.0006,***. To further dissect the effect of the Fc-part of 34-1-2S in TRALI-induction, we aim to investigate the contribution of Fc-mediated complement activation vs Fc-receptor interaction. We therefore successfully generated chimeric variants of 34-1-2S with a humanized IgG1 Fc-domain containing mutations making them functionally complement dead (K322A), Fc-receptor dead (dG236) or both complement and Fc-receptor dead (PG LALA). We are currently investigating the effects of these variants on in vivo TRALI induction.We next investigated how the in vivo murine TRALI reaction was related to numbers of macrophages, monocytes and neutrophils in blood and lungs. We found that 34-1-2S-TRALI was associated with significantly decreased levels of macrophages in the lungs and increased levels in blood, compared to mice infused with PBS, SF1.1.10 or 34-1-2S-Fab, suggesting that Fc-mediated complement activation and TRALI induction is related to macrophage trafficking from lungs to blood.We did not observe any significant differences between blood and lung neutrophil levels of 34-1-2S-TRALI mice compared to SF1.1.10 TRALI-resistant mice. We hypothesized that in TRALI neutrophils undergo formation of neutrophil extracellular traps (NETs) induced by complement. We observed that C5a enabled potent neutrophil-chemotaxis in vitro (P: 0.0048,**). In addition, using direct immunofluorescence staining of extracellular DNA and Citrullinated histone H3, we observed that both LPS and C5a on their own could induce NET formation in vitro, which was synergistically increased with a combination of both LPS and C5a (P: 0.0417,*), as occurring during TRALI. Furthermore, we found increased levels of NETs to be present in plasma samples of TRALI patients (n=53) compared to healthy controls (n=30): 1.64 ± 0.97 vs 0.80 ± 0.34 MPO-DNA OD, respectively, P: 0.0002,***.Finally, we targeted the C5a-receptor (C5aR) using a C5aR-antagonist in our TRALI mouse model. Surprisingly, this did not prevent but even worsened TRALI (P: 0.0398,*), with elevated levels of blood monocytes and macrophages. This suggests that an approach directly targeting complement components may be a more promising therapeutic strategy to explore in combatting TRALI. DisclosuresNo relevant conflicts of interest to declare.

Experimental study on the expression and diagnostic significance of I-FABP in acute intestinal ischemia

Objective: To detect the expression and distribution of I-FABP in intestinal tissue and the changes of serum concentrations at different time of acute intestinal ischemia, and explore the significance and mechanism of I-FABP in early diagnosis of acute ischemic bowel disease. Methods: The selected 96 healthy adult SD rats were randomly divided into the experimental group and control group; 48 in each group. Each group was randomly subdivided into 6 groups with 8 rats in each group. The superior mesenteric artery was ligated in the experimental group and the peritoneal switch operation was performed in the control group. The venous blood samples were extracted from each group rats’ right ventricle at 0.5 h, 1 h, 2 h, 4 h, 8 h, 12 h after the operation and the concentration of I-FABP was tested respectively. Then the rats were killed, and the diseased intestinal tubes were cut out for paraffin sections. The I-FABP in intestinal tissue was stained by routine HE staining and direct immunofluorescence staining. Results: The I-FABP was mainly expressed in the epithelial villi of intestinal mucosa, and there was a small amount of expression in the intestinal submucosa and even the muscularis. Within 1 hour of intestinal ischemia, the number of I-FABP positive granules in the intestine and intestinal cavity increased gradually, and then gradually decreased after 1 hour. The difference has statistically significant between the experimental group and the control group (P &lt; 0.05). The serum I-FABP: In the experimental group, the serum I-FABP concentration began to increase at 0.5 h, and reached a peak at 1 h (290. 24 ± 156.69) μg·L–1, then gradually decreased. Compared with the control group, the difference was statistically significant (P &lt; 0.05). Conclusion: I-FABP usually mainly exists in the epithelial cells of intestinal mucosa. When acute intestinal ischemia occurs, the epithelial cells of intestinal mucosa permeability changes; I-FABP expression rapidly releases to intestinal tissue and intestinal cavity, and is absorbed into the blood. Therefore, I-FABP has certain clinical significance in early diagnosis and treatment of acute intestinal ischemia.

Development of lymphocyte subpopulations in local breed chickens.

Background and Aim:Local breeds of chicken are known to have relatively higher disease resistance to many endemic diseases and diseases that are highly virulent in commercial chickens. This study aimed to address the lymphocyte subpopulations in three constitutive immune system organs (thymus, bursa of Fabricius, and spleen) in 30, 8-week-old, male local breed chickens.Materials and Methods:The T (CD3+) and B lymphocytes (Bu-1+) were identified through one-color, direct immunofluorescent staining of the thymus, bursa, and spleen lymphocytes. Likewise, two-color, direct immunofluorescent staining was performed to identify the CD4- and/or CD8-defined T lymphocytes. The proportions of T and B lymphocytes and CD4- and/or CD8 defined chicken lymphocyte subsets in lymphoid suspensions prepared from the thymus, bursa, and spleen were determined by flow cytometry.Results:CD3+ cells, particularly those positive for CD4+CD8–, were dominant in the thymus, whereas cells expressing the Bu-1 marker were predominant in the bursa of Fabricius. The proportion of T and B cells was almost equal in the spleen, with more cells expressing the CD4–CD8+ marker in the red pulp.Conclusion:These findings indicate that local breeds of chicken could serve as a reliable model for studying the immune system of commercial light chicken breeds, due to the similarity in the presence and the distribution of the immune cells.

The role of cytomegalovirus and interleukin-8 in destabilization of atherosclerotic lesions in humans

Relevance. Currently, the role of persistent infections in the atherogenesis development mechanism is not fully understood. Therefore, its important to analyze the role of viral infection against the background of the pro-inflammatory cytokines expression in atherosclerotic plaque destabilization.&#x0D; The aim of the work was a comparative immunohistochemical study of cytomegalovirus (CMV) and IL-8 in different types of human atherosclerotic lesions during their destabilization.&#x0D; Materials and methods. The study was carried out on 130 autopsy samples of human aorta. CMV was detected by direct immunofluorescent antibody staining. IL-8 was detected by two-stage streptavidin-biotin antibody staining.&#x0D; Results. It has been shown that active detection of both CMV and IL-8 is characteristic of atherogenesis foci of the arterial intima with the most intense immune-inflammatory changes. The obtained results indicate the synergism of the cellular response to CMV and IL-8 in the vascular wall during the destabilization atherosclerotic lesions process.&#x0D; Conclusion. According to the results of the work, it can be concluded that both the presence of CMV in atherosclerotic lesions and the high production of IL-8 play a significant role in the formation of unstable atherosclerotic plaques in the vascular wall.

AB024. A case of bullous systemic lupus erythematosus confined to the oral mucosa

Bullous systemic lupus erythematosus (BSLE) is a rare antibody-mediated blistering disease that presents in association with systemic lupus erythematosus (SLE). Features of BSLE can present as a diagnostic challenge as they often mimic other bullous skin diseases. We present a case of a 45-year-old female with history of celiac disease, Raynaud’s syndrome, chilblains, and Sicca symptoms who presented with 10 months of recurrent vesicles and erosions of the gingiva. Review of symptoms was positive for arthralgias and recurrent photosensitive rashes. Physical examination was notable for multiple 4 mm vesicles with mild surrounding erythema confined to anterior upper gingiva without further mucosal or cutaneous involvement. Two biopsies obtained prior to presentation revealed a mixed inflammatory infiltrate with focal eosinophils, negative for viral cytopathic changes. A biopsy of the maxillary frenum for direct immunofluorescence (DIF) demonstrated a weak, fine linear deposition of IgG and IgA along the basement membrane zone initially suspicious for mucous membrane pemphigoid. Serum ELISA for antibodies against BP180, BP230, and collagen VII were negative. Further lab workup was notable for an ANA of 1:160, anti-Ro60 774 (normal 0–19), anti-Sm 20.7 (normal 0–19), and C4 15.1 (normal 16.3–47.8). The patient was found to meet SLICC criteria (joint disease, chronic cutaneous lupus (chilblains), positive ANA, positive Anti-Sm, and low C4) for SLE diagnosis. The patient was started on dapsone 100 mg by mouth daily with improvement in symptoms. BSLE can affect the oral mucosa and has been described in the absence of anti-collagen VII positivity, but has not been described in the absence of cutaneous bullae. Although atypical, the DIF staining pattern in the context of SLE raises support for a diagnosis of oral BSLE. Teaching point: BSLE can manifest as an oral vesicular eruption and can be differentiated from other oral bullous diseases based on histologic and immunopathologic features

Study of vesicobullous lesions of the skin clinical and histopathology correlation

Introduction: Vesiculobullous diseases are a group of disorders in which primary lesion is a vesicle or a bulla on the skin or mucous membrane or both. Though some of the veisculobullous lesions are characteristics in their appearance and distribution, clinically many a times a definitive diagnosis cannot be made by physical examination alone. Then the dermatologists are forced to resort to histopathology for a definite diagnosis and classification. In the present scenario, even though histopathological study is sufficient in most of the cases, it is often accompanied by immunofluorescence antibody tests for confirmation of the diagnosis and monitoring of the disease. Direct immunofluorescence antibody test is a gold standard to confirm the disease and the indirect test helps to monitor the disease. Objectives: This study was carried out to find out clinical and histopathological correlation in diagnosing the disease. Materials and Methods: In the present study, histopathological evaluated vesiculobullous lesions over the period of 2 years. A total number of 35 cases were encountered. Staining procedures done are haematoxylin – eosin staining, leishmann staining and direct immunofluorescence stain. Results: A total number of 35 cases were encountered which constituted 0.70% of overall biopsy specimen and 12.32% of skin biopsy specimen. The pemphigus vulgaris formed commonest disease encountered accounting for 34.30% of all skin lesions. Bullous pemphigoid formed the second most common disease observed (31.34%) affecting both sexes equally. Conclusions: All though the ‘primary’ vesiculobullous lesions of skin are seen in a small group of people, they have been associated with significant a morbidity and mortality. It is important to distinguish each of these entities and separate them appropriate for management and treatment. Even though clinical examination has its own importance, both histopathological and DIF testing is must, as it confirms the diagnosis and

18660 Increased systemic symptoms in patients with positive direct immunofluorescence of skin biospies with Henoch-Schonlein purpura/IgA vasculitis: A retrospective chart review

Direct immunofluorescence (DIF) of skin biospies is an important tool in the diagnosis of Henoch-Schonlein purpura (HSP)/IgA vasculitis. In a retrospective chart review of IgA positive vasculitis patients seen by dermatology over the previous 5 years, we sought to determine whether presence of systemic symptoms classically seen in HSP correlated with a higher risk of renal disease. A total of 14 out of the 22 patients (63.63%) who underwent DIF staining of a skin biopsy tested positive for IgA. Of the patients that were positive for IgA on biopsy, 64.3% reported gastrointestinal symptoms, 28.6% had musculoskeletal involvement, and 14.3% reported fever.

A Study of Normal Epidermal Melanocyte Distribution

Abstract Objective: To elucidate the epidermal melanocyte distribution according to sex, age, and body part with the goal of providing benchmark data for the diagnosis and therapeutic effect evaluation of pigmentary skin diseases. Methods: Epidermal melanocytes and keratinocytes were assessed using direct immunofluorescence staining, and the melanocyte density and epidermal thickness were calculated. The obtained data were statistically analyzed using SPSS Version 20.0 software. An independent-samples t test was used to compare the data between two groups, while data among three or more groups were compared by one-way analysis of variance. Data correlations were evaluated using Pearson correlation analysis. Results: Melanocytes were uniformly distributed among the keratinocytes in the basal layer, and the average ratio of melanocytes to keratinocytes was 1:7. Among them, the ratio in males was 1:6.5 and that in females was 1:7.4, with no significant difference (P = 0.127). The melanocyte density gradually declined as age increased; the ratio was 1:5.8 before 50 years of age without an obvious downtrend. The average melanocyte density was 1:7.9 within 51 to 65 years of age and 1:8.5 at &gt;65 years of age, and the difference was statistically significant (P &lt; 0.01). Obvious differences were found in the melanocyte density among different body parts; in descending order, these densities were as follows: face (1:4.0) &gt;neck (1:5.1) &gt;hip (1:5.7) &gt;upper limb (1:7.4) &gt;lower limb (1:8.3) &gt;lower back (1:9.2) &gt;thorax and abdomen (1:9.9). The melanocyte density was not related to the epidermal thickness. Conclusion: The melanocyte density showed a declining trend with age and significantly changed after 50 years of age. The melanocyte density was associated with body part; specifically, the density in the face, neck, and hip was higher than that in the limbs and torso. However, the melanocyte density was not associated with sex or epidermal thickness.

S1986 Mucous Membrane Pemphigoid: A Rare Cause of Hematemesis

INTRODUCTION: Mucous membrane pemphigoid (MMP) rarely presents with esophageal involvement [1-3]. We present a case of a 57 year old man who was evaluated for acute onset dysphagia and severe hematemesis. He denied any caustic ingestion. EGD revealed severe esophageal mucosal injury with bleeding, mucosal sloughing with exposure of the submucosal layers, and mid-esophageal narrowing. Biopsies were non-diagnostic. Physical exam revealed erythematous plaques and few bullae on his extremities. Shave biopsies were positive for linear basement membrane (BM) immunofluorescent staining, consistent with MMP. Serum ELISA was positive for Anti-BP180 Ab, which along with the clinical presentation, confirmed the diagnosis of MMP. The patient was started on corticosteroids and mycophenolate mofetil with resolution of his symptoms and evidence of mucosal healing on repeat EGD. CASE DESCRIPTION/METHODS: MMP with esophageal involvement typically presents with longstanding dysphagia due to esophageal strictures that often require dilation. In this atypical case, the patient developed acute dysphagia and hematemesis. EGD revealed an esophageal stricture which was not traversed initially given sloughing mucosa and concern for causing perforation. Fortunately, there was no apparent stricture on repeat EGD done after immunosuppressive therapy was started. Establishing a diagnosis of MMP is essential as treatment requires systemic immunosuppression [1,4]. Esophageal biopsies with direct immunofluorescence (DIF) is the gold standard for diagnosis, but biopsies are frequently inconclusive [2,5]. Therefore, both lesional and perilesional biopsies are recommended for histologic and DIF staining, respectively [2,6]. Indirect immunofluorescence (IIF) and serum ELISA testing can help distinguish MMP from other entities [6]. Anti-BP180 Ab was elevated in our patient and down-trended with immunosuppression. Other more specific IIF tests can help distinguish MMP from bullous pemphigoid. DISCUSSION: MMP involving the esophagus should be considered for patients with unexplained hematemesis or dysphagia, particularly when presenting with mucosal sloughing or strictures. The diagnosis is made with the combination of clinical assessment, histopathology and specialized testing. A high index of suspicion is required to make the diagnosis given that biopsies are often inconclusive. This case stresses the significance of a thorough physical exam and the importance of obtaining lesional and perilesional biopsies to establish a diagnosis of MMP.Figure 1.: Endoscopic Evaluation.