The Free Sign in morphea and other sclerosing disorders: A clue to the presence of early sclerosis?

BACKGROUND: The Floating Sign is a histopathologic clue to the diagnosis of autoimmune sclerosing skin disorders such as morphea and interstitial granulomatous dermatitis (IGD). On the other hand, the "free-floating" sign has been associated with neoplasms, for example, dermatofibroma and interstitial mycosis fungoides. Herein, we report the Free Sign in sclerosing skin disorders. METHODS: In a case-control study, we applied detailed histopathologic definitions of Floating Sign and Free Sign to assess their presence in morphea, IGD, and other sclerosing disorders. RESULTS: Free Sign was present in most cases of morphea (46/55, 84%) and IGD (7/13, 54%) but not necrobiosis lipoidica (NL) (6/14, 42.8%) or sclerodermoid graft versus host disease (SGVHD) (2/7, 28.5%). The sensitivity and specificity of Free Sign for morphea versus other disorders was 84% and 56%, respectively. Floating Sign was not identified in most cases: NL (3/14, 21.4%), SGVHD (1/7, 14.2%), morphea (5/55, 9%), IGD (1/13, 7.7%). The diagnostic sensitivity of Floating Sign in morphea was 9%. CONCLUSIONS: The Free Sign was present in most cases of morphea in our series and may represent a clue to the presence of evolving sclerosis. Free Sign may be seen in other sclerosing disorders. Technical artifact is a potential cause of a false-positive Free Sign.

Functional significance of MAIT cells in psoriatic arthritis.

BACKGROUND: Mucosal-associated invariant T (MAIT) cells are gaining more relevance for autoimmune diseases because of its (i) innate and adaptive immune response (ii) tissue homing properties (iii) production of IL-17A. These cells are predominantly CD8+ cells, because of its strong association with MHC-I. Tc17 CD8+/MAIT cells likely to have a critical role in psoriatic arthritis (PsA). Herein, we have explored pathological significance of MAIT cell in PsA. METHODS: Peripheral blood mononuclear cells (PBMC) and synovial fluid mononuclear cells (SFMC) were collected from age/sex matched (n = 10 for each) PsA, rheumatoid arthritis (RA) and osteoarthritis patients (OA). Hi-D FACS studies were performed: (i) activated memory cells (CD3+CD45RO+) T cells were identified (ii) gating strategies were made to identity the MAIT (CD3+Vα7.2TCR+CD161hi) cells, its phenotype pattern; and functional significance in respect to IL-17A production and responsiveness to human rIL-23. Anti CD3/CD28 ab cocktail was used to activate cells along with rIL-23 to culture and enrich the MAIT cells. The percentages of each cell population and the mean fluorescence intensity (MFI) were analyzed using Flow Jo software. RESULTS: MAIT cells were enriched in synovial fluid of PsA (4.29 ±â€¯0.82%) compared to PBMC (1.04 ±â€¯0.71). With stimulation, SFMC MAIT cells produced significantly more IL-17A (32.66 ±â€¯4.01%) compared to that of RA (23.93 ±â€¯2.81%, p < 0.05) and OA (5.02 ±â€¯0.16%, p < 0.05). MAIT cells were predominantly CD8+ (>80%). Significant upregulation of IL-23R was noted in synovial fluid MAIT cells of PsA (24.97 ±â€¯2.33%, p < 0.001) and RA (21.93 ±â€¯2.29%, p < 0.001) compared to that of OA (2.13 ±â€¯2.29). This IL-23R was functionally active as evidenced by profound mitotic effect in presence of rIL-23. CONCLUSION: MAIT cells are poly functional; produce multiple cytokines (IL-17A, IFN-γ, TNF-α). Here, we demonstrated synovial fluid MAIT cells as a major source of IL-17A and majority of MAIT cells were CD8+. Functionally active IL-23R on these migrated MAIT cells brings a new dimension. They may not need MR1 associated activation rather lesional IL-23 in the synovium can independently regulate these critical Tc17 CD8+ MAIT cells. Thus, these cells likely to be a part of the IL-23/IL-17A cytokine network and play a critical role in the pathogenesis of PsA.

Salmonella Typhimurium Enzymatically Landscapes the Host Intestinal Epithelial Cell (IEC) Surface Glycome to Increase Invasion.

Although gut host-pathogen interactions are glycan-mediated processes, few details are known about the participating structures. Here we employ high-resolution mass spectrometric profiling to comprehensively identify and quantitatively measure the exact modifications of native intestinal epithelial cell surface N-glycans induced by S. typhimurium infection. Sixty minutes postinfection, select sialylated structures showed decreases in terms of total number and abundances. To assess the effect of cell surface mannosylation, we selectively rerouted glycan expression on the host using the alpha-mannosidase inhibitor, kifunensine, toward overexpression of high mannose. Under these conditions, internalization of S. typhimurium significantly increased, demonstrating that bacteria show preference for particular structures. Finally, we developed a novel assay to measure membrane glycoprotein turnover rates, which revealed that glycan modifications occur by bacterial enzyme activity rather than by host-derived restructuring strategies. This study is the first to provide precise structural information on how host N-glycans are altered to support S. typhimurium invasion.

Characteristic Changes in Cell Surface Glycosylation Accompany Intestinal Epithelial Cell (IEC) Differentiation: High Mannose Structures Dominate the Cell Surface Glycome of Undifferentiated Enterocytes.

Changes in cell surface glycosylation occur during the development and differentiation of cells and have been widely correlated with the progression of several diseases. Because of their structural diversity and sensitivity to intra- and extracellular conditions, glycans are an indispensable tool for analyzing cellular transformations. Glycans present on the surface of intestinal epithelial cells (IEC) mediate interactions with billions of native microorganisms, which continuously populate the mammalian gut. A distinct feature of IECs is that they differentiate as they migrate upwards from the crypt base to the villus tip. In this study, nano-LC/ESI QTOF MS profiling was used to characterize the changes in glycosylation that correspond to Caco-2 cell differentiation. As Caco-2 cells differentiate to form a brush border membrane, a decrease in high mannose type glycans and a concurrent increase in fucosylated and sialylated complex/hybrid type glycans were observed. At day 21, when cells appear to be completely differentiated, remodeling of the cell surface glycome ceases. Differential expression of glycans during IEC maturation appears to play a key functional role in regulating the membrane-associated hydrolases and contributes to the mucosal surface innate defense mechanisms. Developing methodologies to rapidly identify changes in IEC surface glycans may lead to a rapid screening approach for a variety of disease states affecting the GI tract.

Spindle Cell Hibernoma of Perinephric Adipose Tissue: A Case Report.

Hibernoma is a relatively uncommon benign neoplasm of brown adipose tissue which usually affects head and neck region, thigh and rarely breast. There are different subtypes of hibernoma with the spindle cell-type is the least common one. Herein, we are reporting a case of this rare spindle cell hibernoma in a location which has not been previously reported: perinephric adipose tissue. This tumor was presented as a renal mass on imaging.

Dysuria and Vaginal Pain, Unusual Manifestations of B-acute Lymphoblastic Lymphoma.

Urinary symptoms are one of the most common reasons for emergency visits in females of pediatric age group and can be associated with various conditions like infections (most common), sexual trauma and rarely neoplastic processes. Here, we report a case of a 7-year-old female who presented in the emergency multiple times with the complaints of urinary symptoms and vaginal pain and was empirically treated with antibiotics and antifungals without symptomatic improvement. Her blood tests, physical examination during this time remained unrevealing. She was then transferred to our institution on her third emergency visit for further evaluation. On imaging studies, she was noted to have expansile lesions on her vertebral body at the L4 and T6 levels with compressive myelopathy with multiple bone and soft tissue lesions throughout her lower extremities. Patient developed saddle anesthesia requiring emergent decompression and biopsy of the epidural mass with the final pathology coming back as B-lymphoblastic leukemia/lymphoma. B-ALL/B-LBL is the most common pediatric hematologic malignancy and usually presents with fever, hepatosplenomegaly, lymphadenopathy, bone pain and bleeding. Occasionally, atypical presentations like bone and joint pain, osteoporosis, palpable paravertebral mass have been described. However, this is the first case report to describe a very unusual and unfamiliar presentation of this disease causing significant diagnostic difficulty resulting in delayed treatment. This case report can aid as a reminder that unusual pain or any nonspecific manifestations in pediatric patients, refractory to common treatment should be investigated with extreme diligence not to miss this neoplastic process.

Diagnosing B-cell acute lymphoblastic leukemia in 2 pediatric patients with recent SARS-CoV-2 infection.

COVID-19 infection is still a mystery in terms of its long-term effect on health and its consequences on hematological disorders. Prior studies including ours have shown the abnormal changes in hematopoietic cells in COVID-19 patients. In this article, we are presenting 2 cases of pediatric B-lymphoblastic leukemia (B-ALL) with a previous history of COVID-19 infection. The first case describes a 22-month-old boy presenting with lymphadenopathy, neutropenia, and anemia with concurrent COVID-19 infection without any evidence of a hematolymphoid neoplasm as per bone marrow and lymph node biopsy. However, he presented after 2 months with bone marrow biopsy confirming B-ALL. The second case is that of a 4-year-old girl presenting with B-ALL who has had asymptomatic COVID-19 infection 5 months before this current presentation. Both the cases had complete resolution of COVID-19 infection during the time of presentation with acute leukemia. There were notably 2 rare findings along the course of the patients' illnesses. First, the unusual plasmacytosis in the marrow during active COVID-19 infection in the first patient and the second, is predilection of development of B-ALL following COVID-19. In both the cases the fluorescence in situ hybridization (FISH) studies showed pathologic alteration of the RUNX1 gene. Overall, there are no literature to support a causal association between acute B-ALL and COVID-19. The diagnosis of B-ALL in these patients after COVID-19 infection may be totally unrelated. However, if we consider Greaves proposed 2-hit model for childhood acute leukemia, that an infectious agent can precipitate development of B-ALL in a genetically susceptible individual. Alteration of the RUNX1 gene in both the patients, opens a door for further exploration of the "second-hit" hypothesis regarding an infectious agent precipitating development of B-ALL in a genetically susceptible individual.

IL-22 induced cell proliferation is regulated by PI3K/Akt/mTOR signaling cascade.

OBJECTIVE: Interleukin 22 (IL-22), a relatively new cytokine has been found to induce significant proliferation of human keratinocytes and fibroblast like synoviocytes (FLS) and thus plays an important role in the pathogenesis of autoimmune diseases like psoriasis and rheumatoid arthritis (RA) which are characterized by hyperproliferation of keratinocytes and FLS respectively. PI3K/Akt/mTOR signaling cascade plays crucial role in cell growth and survival. Therefore our objective was to see the regulatory role of PI3K/Akt/mTOR signaling cascade in IL-22 induced proliferation of keratinocytes and FLS. METHODS: Normal human epidermal keratinocytes (NHEK) and FLS were isolated from skin of healthy volunteer's undergone plastic surgery and synovial tissue of psoriatic arthritis (PsA) and RA patients respectively. IL-22 induced proliferation of NHEK and FLS was measured by MTT assay. Phosphorylation of Akt/mTOR was determined by western blot assay and further confirmed by real time polymerase chain reaction (RT-PCR). RESULTS: We observed that IL-22 induced significant proliferation of NHEK and FLS which was effectively inhibited by dual kinase (PI3K/mTOR) inhibitor, NVP-BEZ235 and specific mTOR inhibitor, Rapamycin. In NHEK and FLS, IL-22 significantly induced phosphorylation of Akt and mTOR which was effectively blocked by Rapamycin and NVP-BEZ235. Further we did RT-PCR in NHEK and found that IL-22 significantly upregulated AKT1 and MTOR gene. CONCLUSION: These results show that IL-22 induced proliferation of NHEK and FLS is dependent on PI3K/Akt/mTOR signaling pathway. This novel observation provides the scope to develop new therapeutics targeting PI3K/Akt/mTOR signaling pathway in autoimmune diseases like psoriasis and rheumatoid arthritis.

Acute Appendicitis with Neuronal Hyperplasia and Swelling: A Novel Histologic Mimic of Appendiceal Goblet Cell Adenocarcinoma and Signet-Ring Cell Adenocarcinoma.

Goblet cell adenocarcinoma and signet-ring cell adenocarcinoma are well-known diagnostic pitfalls of routine appendectomy specimens. Here we present a case of acute appendicitis with prominent neuronal (ganglion cell) hyperplasia and swelling which histologically mimics goblet cell adenocarcinoma and signet-ring cell adenocarcinoma. Attention to the cytologic features of the lesional cells (absence of atypia, mitotic activity) and their close association with nerves and classic ganglion cells, along with the use of a limited panel of immunostains, ensures proper classification of this rare but striking benign process.

Leukoerythroblastosis in a Sickle Cell Patient With Pregnancy: An Interesting Peripheral Blood Smear Finding.

The global pandemic of coronavirus disease 2019 (COVID-19) caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) has shaken the entire world. The social, health and financial impacts of this pandemic are beyond words. We have learnt a lot about this new disease in a short period of time, but still a long road to go to fully determine its pathogenic effect. The primary target of this virus is angiotensin-converting enzyme 2 (ACE2) receptor, which is prevalent in endothelial cells throughout the body. Immunocompromised patients such as patients with sickle cell disease are more vulnerable to severe respiratory infections, including infection with SARS-CoV-2. In addition, sickle cell disease patients are prone to vaso-occlusive crisis, and theoretically SARS-CoV-2 can worsen the situation as it also can cause endothelial dysfunction and thrombosis. Herein, we are sharing an interesting peripheral blood smear finding of an asymptomatic 31-year-old multigravida pregnant female with a history of sickle cell disease and found to have a positive COVID-19 polymerase chain reaction (PCR) test during her third trimester of pregnancy at a routine clinic visit. Two weeks after the initial positive test, she developed nausea, vomiting, constipation and a pain crisis affecting her extremities while her COVID-19 PCR test was still positive. She was hemodynamically stable, and lab workup revealed chronic anemia, leukocytosis with neutrophilia and lymphopenia. Morphologic examination of the peripheral blood smear showed a marked leukoerythroblastosis: rare myeloblasts, sickle cells, markedly abundant nucleated red blood cells (RBCs), metamyelocytes, and many large and giant platelets were seen. In this context, her previous peripheral blood smears (prior to positive COVID-19 test) did not show leukoerythroblastosis. She was managed conservatively with hydration and pain control and delivered at 36 weeks via cesarean section due to pre-term labor and intrauterine growth retardation. The unusual finding of leukoerythroblastosis in a pregnant sickle cell disease patient with an asymptomatic COVID-19 infection indicates further studies to determine its effect on hematopoietic system and elucidate its clinical significance.

Immunotherapy in synchronous MSI-H rectal adenocarcinoma and upper tract urothelial carcinoma: a case report.

Background: A growing body of evidence suggests that conventional chemotherapy may not be effective in mismatch repair deficiency (dMMR)/microsatellite instability-high (MSI-H) locally advanced rectal cancer (LARC). Alternative strategies, such as immunotherapy, are currently being investigated both in the neoadjuvant and adjuvant setting. Furthermore, immunotherapy is an attractive alternative to the use of combination chemotherapy regimens when treating synchronous primary cancers such as in the setting of inherited cancer syndromes. Case Description: Here we present a case of a middle-aged woman diagnosed with dMMR/MSI-H locally advanced rectal cancer with synchronous upper tract urothelial cancer secondary to Lynch syndrome. The patient was first treated using neoadjuvant chemotherapy followed by chemoradiation, resulting in only a partial pathologic response. Following surgery, the patient was treated with adjuvant combination immunotherapy with nivolumab, a PD-1 inhibitor, and ipilimumab, a CTLA-4 inhibitor, resulting in a durable disease-free interval of nearly 21 months. Conclusions: This case report illustrates the importance of determining dMMR/MSI-H status in LARC and the consideration of immunotherapy (particularly with synchronous primaries as seen in inherited cancer syndromes), reviews the current literature, and calls for further investigation into the use of neoadjuvant and adjuvant immunotherapy in locally advanced rectal cancer along with upper tract urothelial carcinoma (UTUC).

Effects of Image Quantity and Image Source Variation on Machine Learning Histology Differential Diagnosis Models.

AIMS: Histology, the microscopic study of normal tissues, is a crucial element of most medical curricula. Learning tools focused on histology are very important to learners who seek diagnostic competency within this important diagnostic arena. Recent developments in machine learning (ML) suggest that certain ML tools may be able to benefit this histology learning platform. Here, we aim to explore how one such tool based on a convolutional neural network, can be used to build a generalizable multi-classification model capable of classifying microscopic images of human tissue samples with the ultimate goal of providing a differential diagnosis (a list of look-alikes) for each entity. METHODS: We obtained three institutional training datasets and one generalizability test dataset, each containing images of histologic tissues in 38 categories. Models were trained on data from single institutions, low quantity combinations of multiple institutions, and high quantity combinations of multiple institutions. Models were tested against withheld validation data, external institutional data, and generalizability test images obtained from Google image search. Performance was measured with macro and micro accuracy, sensitivity, specificity, and f1-score. RESULTS: In this study, we were able to show that such a model's generalizability is dependent on both the training data source variety and the total number of training images used. Models which were trained on 760 images from only a single institution performed well on withheld internal data but poorly on external data (lower generalizability). Increasing data source diversity improved generalizability, even when decreasing data quantity: models trained on 684 images, but from three sources improved generalization accuracy between 4.05% and 18.59%. Maintaining this diversity and increasing the quantity of training images to 2280 further improved generalization accuracy between 16.51% and 32.79%. CONCLUSIONS: This pilot study highlights the significance of data diversity within such studies. As expected, optimal models are those that incorporate both diversity and quantity into their platforms.s.

Significance of Pulmonary Hypertension in Hypertrophic Cardiomyopathy.

Hypertrophic cardiomyopathy (HCM) is the most prevalent hereditary cardiac disease characterized by the presence of left ventricular and/or septal hypertrophy in the absence of other underlying cardiac disorders. Patients of HCM have a broad range of clinical presentation from being asymptomatic to severely ill condition requires hospitalization and urgent management. Broadly, HCM is classified in two variants: obstructive and nonobstructive. The mainstay of diagnosis is through echocardiography. As HCM chiefly affect the left heart, pulmonary hypertension (PH) is an expected complication of this disease. Though the existence of PH in HCM is known for a long time, its clinical significance, underlying mechanism, and prognostic impact in HCM have been revealed by few recent studies. Specifically, studies have shown increased events of thromboembolism, atrial fibrillation, and heart failure in patients with HCM and PH. These studies elucidated the underlying mechanism of PH in HCM--a rise of pressure in the precapillary and postcapillary pulmonary vasculature. In addition to left ventricular involvement, studies have shown right ventricular involvement and the association of left and right ventricular dysfunction in these patients. Further, it has been shown that surgical intervention to reduce septal thickness improves survival in pharmacotherapy nonresponders and the presence of PH does not increase mortality in these patients. We present a comprehensive review exploring the prevalence, underlying mechanisms, and impact of PH on HCM.

Limited Number of Cases May Yield Generalizable Models, a Proof of Concept in Deep Learning for Colon Histology.

BACKGROUND: Little is known about the effect of a minimum number of slides required in generating image datasets used to build generalizable machine-learning (ML) models. In addition, the assumption within deep learning is that the increased number of training images will always enhance accuracy and that the initial validation accuracy of the models correlates well with their generalizability. In this pilot study, we have been able to test the above assumptions to gain a better understanding of such platforms, especially when data resources are limited. METHODS: Using 10 colon histology slides (5 carcinoma and 5 benign), we were able to acquire 1000 partially overlapping images (Dataset A) that were then trained and tested on three convolutional neural networks (CNNs), ResNet50, AlexNet, and SqueezeNet, to build a large number of unique models for a simple task of classifying colon histopathology into benign and malignant. Different quantities of images (10-1000) from Dataset A were used to construct >200 unique CNN models whose performances were individually assessed. The performance of these models was initially assessed using 20% of Dataset A's images (not included in the training phase) to acquire their initial validation accuracy (internal accuracy) followed by their generalization accuracy on Dataset B (a very distinct secondary test set acquired from public domain online sources). RESULTS: All CNNs showed similar peak internal accuracies (>97%) from the Dataset A test set. Peak accuracies for the external novel test set (Dataset B), an assessment of the ability to generalize, showed marked variation (ResNet50: 98%; AlexNet: 92%; and SqueezeNet: 80%). The models with the highest accuracy were not generated using the largest training sets. Further, a model's internal accuracy did not always correlate with its generalization accuracy. The results were obtained using an optimized number of cases and controls. CONCLUSIONS: Increasing the number of images in a training set does not always improve model accuracy, and significant numbers of cases may not always be needed for generalization, especially for simple tasks. Different CNNs reach peak accuracy with different training set sizes. Further studies are required to evaluate the above findings in more complex ML models prior to using such ancillary tools in clinical settings.

Optical fiber nanoprobe preparation for near-field optical microscopy by chemical etching under surface tension and capillary action.

We propose a technique of chemical etching for fabrication of near perfect optical fiber nanoprobe (NNP). It uses photosensitive single mode optical fiber to etch in hydro fluoric (HF) acid solution. The difference in etching rate for cladding and photosensitive core in HF acid solution creates capillary ring along core-cladding boundary under a given condition. The capillary ring is filled with acid solution due to surface tension and capillary action. Finally it creates near perfect symmetric tip at the apex of the fiber as the height of the acid level in capillary ring decreases while width of the ring increases with continuous etching. Typical tip features are short taper length (approximately 4 microm), large cone angle (approximately 38 degrees ), and small probe tip dimension (<100 nm). A finite difference time domain (FDTD) analysis is also presented to compare near field optics of the NNP with conventional nanoprobe (CNP). The probe may be ideal for near field optical imaging and sensor applications.

Correction to: Inflammation, Atherosclerosis, and Psoriasis.

Following publication of this article [1] it came to our attention that we neglected to reference an article by Ishwarlal Jialal published in Critical Pathways in Cardiology [2].

Classic Ulcerative Pyoderma Gangrenosum Is a T Cell-Mediated Disease Targeting Follicular Adnexal Structures: A Hypothesis Based on Molecular and Clinicopathologic Studies.

BACKGROUND: Pyoderma gangrenosum (PG) is a debilitating ulcerative skin disease that is one of the most common associated diseases seen in patients with inflammatory bowel disease and rheumatoid arthritis. Although PG is classified as a neutrophilic dermatosis, its pathophysiology is poorly understood. OBJECTIVE: Use data obtained from patient-reported histories, immunohistochemistry, and gene expression analysis to formulate a hypothesis on PG pathophysiology. METHODS: Ten PG patients participated and answered questions about new ulcer formation. Skin biopsies of healed prior ulcers and adjacent normal skin were obtained from four patients for immunohistochemistry. Scars from healthy patients and patients with discoid lupus were used as additional controls. New onset PG papules were analyzed using immunohistochemistry and gene expression analysis via quantitative real-time PCR. RESULTS: All PG patients reported that healed sites of previous ulceration are refractory to re-ulceration. Simultaneous biopsies of healed and uninvolved skin triggered ulceration only in the latter. On immunohistochemistry, healed PG scars showed complete loss of pilosebaceous units, which were present in normal skin, and to a lesser extent in control scars, and discoid scars. Early PG papules showed perivascular and peripilosebaceous T cell infiltrates, rather than neutrophils. These early inflammatory events were dominated by increased gene expression of CXCL9, CXCL10, CXCL11, IL-8, IL-17, IFNG, and IL-36G and transcription factors consistent with Th1 phenotype. LIMITATIONS: Small sample size was the main limitation. CONCLUSION: We put forth the hypothesis that PG is a T cell response resulting in the destruction of pilosebaceous units.