T follicular helper cells expansion in transplant recipients correlates with graft infiltration and adverse outcomes.

Introduction: The process of immunization following vaccination in humans bears similarities to that of immunization with allografts. Whereas vaccination aims to elicit a rapid response, in the transplant recipient, immunosuppressants slow the immunization to alloantigens. The induction of CD4+CXCR5+ T follicular helper (Tfh) cells has been shown to correlate with the success of vaccine immunization. Method: We studied a cohort of 65 transplant recipients who underwent histological evaluation concurrent with PBMC isolation and follow-up sampling to investigate the phenotypic profiles in the blood and allotissue and analyze their association with clinical events. Results: The proportion of circulating Tfh cells was heterogeneous over time. Patients in whom this compartment increased had lower CCR7-PD1+CD4+CXCR5+ T cells during follow-up. These patients exhibited more alloreactive CD4+ T cells using HLA-DR-specific tetramers and a greater proportion of detectable circulating plasmablasts than the controls. Examination of baseline biopsies revealed that expansion of the circulating Tfh compartment did not follow prior intragraft leukocyte infiltration. However, multicolor immunofluorescence microscopy of the grafts showed a greater proportion of CXCR5+ T cells than in the controls. CD4+CXCR5+ cells were predominantly PD1+ and were in close contact with B cells in situ. Despite clinical stability at baseline, circulating Tfh expansion was associated with a higher risk of a composite of anti-HLA donor-specific antibodies, rejection, lower graft function, or graft loss. Conclusion: In otherwise stable patients post-transplant, circulating Tfh expansion can identify ongoing alloreactivity, detectable before allograft injury. Tfh expansion is relevant clinically because it predicts poor graft prognosis. These findings have implications for immune surveillance.

Chromophobe Renal Cell Carcinoma with Sarcomatoid Differentiation: Clinicopathologic Correlation and Molecular Findings.

Introduction: Sarcomatoid differentiation has been reported in approximately 8% of chromophobe renal cell carcinoma (RCC) and is associated with a worse prognosis. We aim to describe the clinicopathologic and molecular findings of chromophobe RCC with sarcomatoid differentiation. Methods: Surgical pathology database was searched to identify chromophobe RCC with sarcomatoid differentiation from January 2015 to December 2021. Results: Five patients were diagnosed with chromophobe RCC with sarcomatoid differentiation. The median age at the time of diagnosis was 57 years (range 51-61 years). Three patients died after median follow-up of 12.1 months (range 1.6-18.2 months). The median tumor size was 10.7 cm (range 5.6-13.6 cm). The median percentage of sarcomatoid component was 60% (range 10-90%), and the median percentage of necrosis was 30% (range 10-50%). One tumor demonstrated osteoid formation. PAX8, keratin 7, KIT (CD117), and Hale colloidal iron were positive in the epithelial component, whereas the sarcomatoid component was positive for vimentin, CD10, and high Ki67 proliferative index. Molecular testing was performed in three specimens: all were TP53 mutated and microsatellite stable. One aggressive tumor had RB1 frameshift mutation and copy number gains for TERT and CUL4A. Conclusion: Chromophobe RCC with sarcomatoid differentiation is a rare entity with aggressive behavior. Percentage of sarcomatoid component, necrosis, and the occurrence of metastasis is associated with worse prognosis. Molecular profiling reveals frequent TP53 mutation. While TERT promoter mutation has no prognostic implication, FLCN inactivation may be associated with a less aggressive course. The clinical significance of RB1 loss is unclear.

From principles to practice: Implementation of entrustable professional activities (EPAs) for surgical pathology residency education in a large academic hospital.

Over the past decade, competency-based medical education (CBME) has gained momentum in the United States to develop trainees into independent and confident physicians by the end of their training. Entrustable professional activities (EPAs) are an established methodology for assessing trainee development through an outcomes-driven rather than a time-based model. While EPAs have been utilized as an assessment tool for CBME in Europe and Canada, their validation and implementation in some medical specialties has occurred more recently in the United States. Pediatrics was the first specialty in the US to conduct a large-scale UME-GME pilot. Pathology Residency EPAs were published in 2018; however, implementation in training programs has been slow. We have piloted EPAs in our residency program's surgical pathology rotation and propose a unique set of 4 surgical pathology EPAs to track trainee preparedness for independent practice.

A novel 6-day cycle surgical pathology rotation improves resident satisfaction and maintains Accreditation Council for Graduate Medical Education (ACGME) milestone performance.

Surgical pathology residency training in the United States lags behind other specialties in quality control and graduated responsibility to train independent pathologists capable of seamlessly entering practice after training. We observed that our traditional 3-day-cycle surgical pathology cycle (day 1-grossing; day 2 -biopsies/frozens/preview; day 3 - sign-out) consistently and negatively impacted resident education by reducing preview time, case follow-up, immunohistochemical stain (IHC) interpretation, and molecular study integration. We aimed to create a modern surgical pathology rotation that improved performance and outcomes. We innovated our rotation to enhance resident education and ensure graduated responsibility. A novel 6-day cycle was created composed of 2 grossing days, 1 frozens/biopsies/preview days, 2 dedicated sign-out days, and 1 frozens/biopsies/case completion day. Residents completed surveys before implementing the new rotation and 6 months after implementation to track self-assessment of Accreditation Council for Graduate Medical Education (ACGME) milestone performance and internal quality control metrics. Clinical Competency Committee (CCC) annual evaluations were assessed in paired PGY levels pre- and post-intervention. After implementation, there was a statistically significant improvement in self-assessment of levels 4 and 5 of ACGME milestones and improved satisfaction of quality metrics, including time for previewing, reviewing IHC, graduated responsibility, and perceived readiness for independent practice. CCC evaluations showed overall maintained performance levels, with trends towards improvements in junior resident classes. Our 6-day cycle adequately fulfills the current demands of our sizeable academic center's surgical pathology training and can be a model for pathology residencies looking to modernize their surgical pathology rotations and resident education.

Alloimmunization Against RBC Antigens Is Not Associated With Decreased Survival in Liver Transplant Recipients.

BACKGROUND: Improvement of liver transplantation (LT) outcomes requires better understanding of factors affecting survival. The presence of RBC alloantibodies (RBCAs) on survival in LT recipients was evaluated. METHODS: This study was a single-center, retrospective cohort study reviewing transfusion records and all-cause mortality between 2002 and 2021. RESULTS: Between 2002 and 2021, 2079 LTs were completed, 1,396 of which met inclusion criteria (1,305 RBCA negative; 91 RBCA positive [6.5%]). The cohorts were similar in age (mean [range], 55.8 [17-79] years vs 56.8 [25-73] years; P = .41, respectively) or sex (RBCA negative, 859 [65%] men and 446 [35%] women vs RBCA positive, 51 [56%] men and 40 [44%] women; P = .0684). Of 132 RBCAs detected, 10 were most common were to E (27.27%), Jka (15.91%), K (9.09%), C (8.33%), M (6.06%), D (5.3%), Fya (4.55%), e (2.27%), c (2.27%), and Jkb (2.27%). Twenty-seven patients (29.7%) had more than 1 RBCA; the most common combinations were C with Jka (7.4%) and E with Dia (7.4%). All-cause mortality was increased in men (men, 14.45 years vs women, 17.27 years; P = .0266) and patients 65 years of age and older (≥65 years of age, 10.21 years vs <64 years of age, 17.22 years; P < .0001). The presence of RBCA (≥1) did not affect all-cause mortality (RBCA negative, 14.17 years vs RBCA positive, 15.29 years; P = .4367). The top 5 causes of death were infection (11.9%), primary malignancy (solid) (10.8%), recurrent malignancy (10.5%), cardiovascular arrest (7.1%), and pulmonary insufficiency/respiratory failure (5.7%). CONCLUSIONS: Survival in RBCA-positive LT recipients is no different from that in RBCA-negative LT recipients.

Cytomorphology of renal angiomyolipoma: performance and accuracy of touch preparation of core needle biopsy.

INTRODUCTION: The recognition of renal angiomyolipoma (AML) can be challenging based on cytology preparations such as touch preparation (TP) of core needle biopsy (CNB) and fine needle aspiration. This study evaluated the cytologic features and performance of TP of CNB during rapid onsite evaluation (ROSE) of renal AML. MATERIALS AND METHODS: A pathology database search was performed between 2000 and 2021 for renal CNB specimens with ROSE using TP that were primarily favored AML on preliminary impression and/or confirmed AML on CNB or subsequent resection. RESULTS: Twenty confirmed AML were identified (90% female, median age 65.5 years). Sixteen (80%) were deemed adequate for diagnosis at the time of ROSE, and 9 of 16 (56%) had available onsite impression: AML was favored in 4 of 9 cases (44%). Examination of TP slides revealed spindle/epithelioid cells in 20 (100%), adipose tissue in 14 (70%), and blood vessels in 3 (15%). All AML cases were subsequently confirmed by immunohistochemistry. Additionally, 3 other cases with ROSE favoring AML revealed to be "renal parenchyma with fibrosis," clear cell papillary renal cell tumor and clear cell renal cell carcinoma. CONCLUSIONS: Onsite evaluation of TP ensures adequate material for diagnosis in most renal AML. Spindle/epithelioid cells were the most common component seen on TP, followed by adipose tissue. Blood vessels were rarely seen. While the recognition of AML at ROSE can be challenging, proper evaluation is important in obtaining adequate diagnostic tissue. Correlation with CNB and utilization of immunohistochemistry are crucial for arriving at the diagnosis.

Minimally invasive cytologic evaluation leading to the diagnosis of TFE3-rearranged renal cell carcinoma: A case report.

TFE3-rearranged renal cell carcinoma (RCC) has been categorized as a molecularly defined renal carcinoma in the 2022 WHO classification of tumors as it does not demonstrate a specific genotype-phenotype correlation. However, in order to arrive at the diagnosis, recognition of the broad spectrum of cytologic and histologic features that can be seen in TFE3-rearranged RCC is important for differential diagnostic consideration. Reported here is the diagnostic workup of a TFE3-rearranged RCC using very limited tissue sample. The initial evaluation was dependent on the cytomorphologic findings observed on a touch preparation made from the renal mass biopsy, directing appropriate selection of ancillary tests, and leading to a definitive diagnosis.

Endothelial colony forming cells generated from cryopreserved peripheral blood mononuclear cells.

Derivation of endothelial colony forming cells (ECFCs) from peripheral blood mononuclear cells (PBMCs) is a technique that could provide access to donor endothelial cells to study donor endothelium/recipient immune cells interactions. The success rate of ECFC colony formation from cryopreserved PBMCs has not been reported. We used biobanked PBMCs and studied the yield of ECFC generation. Endothelial phenotype was confirmed with CD31, CD146, CD309, CD34, CD14 and CD11c staining by flow cytometry and VE-cadherin, von Willebrand factor and Dil-Ac-LDL by fluorescent microscopy. Functionality was tested by endothelial cell tube-based formation assay. The success rate of ECFC generation was 28%. Freezing time was not a predictor of ECFC generation while a shorter time on dialysis and living transplant were significant determinants. These data suggest that it is possible to generate ECFCs from cryopreserved PBMCs, which is a potentially useful option for the longitudinal assessment of alloimmune response in transplantation.

Longitudinal immune profile reveals reduced function of pro-inflammatory monocytes with age following kidney transplantation.

Toxicity of immunosuppression, notably the risk of infection, increases with age. However, the dynamic changes in innate immune response following transplantation are unclear. Based on recent observations, we hypothesized that pro-inflammatory capacity would decrease with age. We analyzed approximately 300 PBMC samples collected longitudinally in 45 de novo, adult kidney recipients and performed detailed phenotypic and functional profiling of monocytes and T cell subsets. Inflammatory response to TLR4 stimulation and indirect allostimulation using mismatched HLA peptides were assessed. In patients aged ≥56 years, TNF-α production by intermediate monocytes was similar to that in younger patients early posttransplant, but diminished substantially later. Adjusted analyses suggested that this was not attributable to confounding factors. In contrast, the alloimmune response to HLA peptides measured by IFN-γ in CD4+ T cells and TNF-α in monocytes was stable over time, but was low in older recipients. Measurement of CD80-86 surface expression revealed no signal for a lower costimulation capacity of APCs. These results suggest that older recipients have a reduced function of their innate pro-inflammatory immune cells posttransplant while maintaining a stable, low alloimmune response over time. The effect of reduced immunosuppressant doses on preventing this phenomenon needs to be clarified.

A Sequential Two-Step Cell-Based Assay Predicts Immunosuppression-Related Adverse Events.

Immunosuppressants are associated with serious and often life-threatening adverse effects. To optimize immunotherapy, a tool that measures the immune reserve is necessary. We validated that a cell-based assay that measures TNF-α production by CD14+16+ intermediate monocytes following stimulation with EBV peptides has high sensitivity for the detection of over-immunosuppression (OIS) events. To develop a sequential, two-step assay with high specificity, we used PBMCs from kidney recipients (n = 87). Patients were classified as cases or controls, according to the occurrence of opportunistic infection, recurring bacterial infections, or de novo neoplasia. Patients who tested positive in the first step were randomly allocated to a training or a testing set for the development of the second step. In the discovery phase, an assay based on the examination of early mature B (eBm5) cells was able to discriminate OIS patients from controls with a specificity of 88%. The testing set also revealed a specificity of 88%. The interassay coefficient of variability between the experiments was 6.1%. Stratified analyses showed good diagnostic accuracy across tertiles of age and time posttransplant. In the adjusted model, the risk of OIS was more than 12 times higher in patients classified as positive than in those who tested negative (adjusted hazard ratio, 12.2; 95% confidence interval: 4.3-34.6). This sequential cell-based assay, which examines the monocyte and eBm5 cell response to EBV peptides, may be useful for identifying OIS in immunosuppressed patients.

C-reactive protein as an inflammatory marker of acute infections outside intensive care settings: case report and evidence-based literature review.

We present the clinical case of a 76-year-old woman with delirium caused by multiple factors, including pneumonia. Although this type of case is quite common in clinical practice, it provides us with an opportunity to discuss laboratory testing in this context, with a special focus on the role of C-reactive protein (CRP). We present data regarding the requests for determination of serum CRP levels at the University of São Paulo University Hospital over the past few years. We also present a review of the medical literature on the topic, as well as clinical epidemiology concepts related to the impact that CRP testing has on the medical decision-making process.