Validating Reference Gene Expression Stability in Human Ovarian Follicles, Oocytes, Cumulus Cells, Ovarian Medulla, and Ovarian Cortex Tissue

Human ovarian cells are phenotypically very different and are often only available in limited amounts. Despite the fact that reference gene (RG) expression stability has been validated in oocytes and other ovarian cells from several animal species, the suitability of a single universal RG in the different human ovarian cells and tissues has not been determined. The present study aimed to validate the expression stability of five of the most used RGs in human oocytes, cumulus cells, preantral follicles, ovarian medulla, and ovarian cortex tissue. The selected genes were glyceraldehyde 3-phosphate dehydrogenase (GAPDH), beta-2-microglobulin (B2M), large ribosomal protein P0 (RPLP0), beta-actin (ACTB), and peptidylprolyl isomerase A (PPIA). Overall, the stability of all RGs differed among ovarian cell types and tissues. NormFinder identified ACTB as the best RG for oocytes and cumulus cells, and B2M for medulla tissue and isolated follicles. The combination of two RGs only marginally increased the stability, indicating that using a single validated RG would be sufficient when the available testing material is limited. For the ovarian cortex, depending on culture conditions, GAPDH or ACTB were found to be the most stable genes. Our results highlight the importance of assessing RGs for each cell type or tissue when performing RT-qPCR analysis.

Acceleration of amyloid fibril formation by multichannel sonochemical reactor

Formation of amyloid fibrils of various amyloidogenic proteins is dramatically enhanced by ultrasound irradiation. For applying this phenomenon to the study of protein aggregation science and diagnosis of neurodegenerative diseases, a multichannel ultrasound irradiation system with individually adjustable ultrasound-irradiation conditions is necessary. Here, we develop a sonochemical reaction system, where an ultrasonic transducer is placed in each well of a 96-well microplate to perform ultrasonic irradiation of sample solutions under various conditions with high reproducibility, and applied it for studying amyloid-fibril formation of amyloid $\beta$, $\alpha$-synuclein, $\beta$2-microglobulin, and lysozyme. The results clearly show that our instrument is superior to conventional shaking method in terms of degree of acceleration and reproducibility of fibril formation reaction. The acceleration degree is controllable by controlling the driving voltage applied to each transducer. We have thus succeeded in developing a useful tool for the study of amyloid fibril formation in various proteins.

Urinary Beta-2-Microglobulin and Late Nephrotoxicity in Childhood Cancer Survivors

The objectives of this study were to evaluate urinary beta-2-microglobulin (β2M) levels in long-term childhood cancer survivors and to establish its association with anticancer drug-induced nephrotoxicity. The study consisted of 165 childhood cancer survivors (CCS) who were in continuous complete remission. We reported that CCS had a significantly higher level of β2M (p < 0.001) and β2M/Cr. ratio (p < 0.05) than healthy peers. Among all participants, 24 (14.5%) had decreased eGFR (<90 mL/min/1.73 m2). A significant positive correlation between β2M/Cr. ratio and body mass index (coef. 14.48, p = 0.046) was found. Furthermore, higher levels of urinary β2M were detected among CCS with a longer follow-up time (over 5 years) after treatment. Subjects with decreased eGFR showed statistically higher urinary β2M levels (20.06 ± 21.56 ng/mL vs. 8.55 ± 3.65 ng/mL, p = 0.007) compared with the healthy peers. Twelve survivors (7.2%) presented hyperfiltration and they had higher urinary β2M levels than CCS with normal glomerular filtration (46.33 ± 93.11 vs. 8.55 ± 3.65 ng/mL, p = 0.029). This study did not reveal an association between potential treatment-related risk factors such as chemotherapy, surgery, radiotherapy, and the urinary β2M level. The relationship between treatment with abdominal radiotherapy and reduced eGFR was confirmed (p < 0.05). We demonstrated that urinary beta-2-microglobulin may play a role in the subtle kidney injury in childhood cancer survivors; however, the treatment-related factors affecting the β2M level remain unknown. Further prospective studies with a longer follow-up time are needed to confirm the utility of urinary β2M and its role as a non-invasive biomarker of renal dysfunction.

Racial Disparities in the Diagnostic Evaluation of Multiple Myeloma

Introduction: Multiple myeloma (MM) is the most common hematologic malignancy in Black individuals with a 2-to-3-fold higher incidence rate among Black compared to White individuals. While therapeutic advances have led to significant increases in survival rates in MM across races, there still exist racial disparities in outcomes that have been attributed to inferior access to novel therapies and autologous stem cell transplant among Black patients. Risk stratification is an important strategy that allows clinicians to identify high-risk disease and potentially tailor therapy based on staging to try to abrogate its poor prognosis. Current risk stratification schemata in MM, such as the International Staging System (ISS) and Revised ISS (R-ISS), necessitate serum laboratory data and fluorescence in-situ hybridization (FISH) cytogenetic analysis of bone marrow specimens. To the best of our knowledge, it is unknown if discrepancies exist in the initial diagnostic workup of MM between Blacks and Whites, which ultimately may have treatment and outcome implications. We sought to assess the presence of racial disparities in the diagnostic workup of newly diagnosed MM among Black and White patients. Methods: We performed a retrospective cohort study using the Surveillance, Epidemiology and End Results (SEER)-Medicare linked database, which includes 16,174 MM patients with patient-level demographics, survival data, and health care claims information. The data included patients ≥ 65 years-old with the diagnosis of MM between 2001-2015. Race was documented by SEER registries. Lab and imaging data were collected from 180 days before and after diagnosis to capture an ample and appropriate allotted time for diagnostic workup. CPT codes were queried to determine the frequency of the diagnostic tests of interest. Data were analyzed through R version 4.0.2. Pearson chi-squared tests were used to compare frequency of diagnostic testing between racial groups. Results: A total of 18,267 MM patients were identified in the SEER-Medicare linked database. Of that, 15,360 MM patients (12,645 White and 2,715 Black) were identified with peripheral blood laboratory, bone marrow, and imaging health care claims data available . The remaining 2,907 patients with a documented race other than White or Black were excluded. Complete blood count and comprehensive metabolic panel serum tests were used to evaluate completeness of CPT codes use, which demonstrated that &gt;89% (13,723/15,360) of individuals had both tests performed. Overall, Black patients had lower frequency of nearly all serum and imaging tests completed relative to White patients (Table 1). Only 61% of White patients underwent the testing components necessary to adequately risk-stratify disease by ISS (e.g., beta-2 microglobulin) compared to 50% of Black patients (relative difference 21%). 30% of White individuals underwent the components to determine R-ISS (e.g., FISH cytogenetics). There were low overall rates of FISH cytogenetics obtained in the study population, with White individuals undergoing FISH cytogenetics at a rate of 30% compared to 25% among Black individuals (relative difference 18%). Magnetic resonance imaging (MRI) of the lumbar spine, the most commonly imaged portion of the spine by MRI, was performed more commonly in White vs Black individuals (33% vs 24%, relative difference 35%). PET/CT was performed in only a small percentage of patients (Whites 9% vs Blacks 5%, relative difference 94%). Conclusions: In a real-world analysis of patients with newly diagnosed MM, we found that Black patients were less likely than White patients to undergo a complete initial diagnostic evaluation. While rates of beta-2 microglobulin and FISH cytogenetics testing were low among all patients, Black patients were less likely to undergo testing needed to complete staging for ISS/R-ISS or proper imaging to assess for extramedullary disease (i.e., PET/CT). Whether these differences between the races in the initial diagnostic workup lead to differences in treatment strategies and survival outcomes deserves additional investigation. Further work is needed to improve access to complete diagnostic evaluation among Black patients with newly diagnosed MM. Figure 1 Figure 1. Disclosures Calip: Pfizer: Research Funding; Roche: Current equity holder in publicly-traded company; Flatiron Health: Current Employment. Derman: Sanofi: Membership on an entity's Board of Directors or advisory committees.

Biological and Classical Prognostic Factors with Impact on Overall Survival in Patients with Early Stage (I/II) Diffuse Large B-Cell Lymphoma: A Study from the Grupo De Estudio Latinoamericano De Linfoproliferativos (GELL)

Background: Diffuse large B-cell lymphoma (DLBCL) is the most common non-Hodgkin lymphoma subtype. Early stage (I/II) disease is seen in up to 30% of all DLBCL cases, and although outcomes in this subgroup have been reported as optimal, relapses can still occur. Prognostic models such as the International Prognostic Index (Miller, NEJM, 1998) continues to be utilized for risk stratification in DLBCL. However, despite its limitations and lack of validation in specific demographic groups such as Latin American patients, no prognostic models exist for the evaluation of limited stage DLBCL. Therefore, we aim to investigate different clinico-epidemiological and laboratory variables and its impact on survival in early stage DLBCL. Methods: We conducted a retrospective study of newly diagnosed patients with early stage DLBCL. Using the Grupo de Estudio Latinoamericano de Linfroproliferativos (GELL) database, we selected patients that had early stage disease, defined as non-bulky stage I or II. The variables analyzed included demographic and clinical variables (e.g., age, ECOG performance status), the International Prognostic Index (IPI), and laboratory variables such as serum albumin, serum lactate dehydrogenase (LDH), serum beta-2-microglobulin, the lymphocyte-monocyte ratio (LMR), the neutrophil-lymphocyte ratio (LNR), and the platelet-lymphocyte ratio (PLR). To determine the variables associated with mortality, univariate and multivariate Cox regression (step-wise type) analysis was performed. Kaplan-Meier and log-rank test were used for survival analysis. Outcomes with a p-value &lt;0.05 were considered statistically significant. Results: We identified 1,375 patients with DLBCL; 503 were identified as early stage DLBCL of whom 498 had sufficient data for analysis. Almost all cases (n=483, 96%) had nodal disease as the primary site, and 15 (4%) extranodal, all within the gastrointestinal tract. There was a slight female predominance (51.8%). The median age at diagnosis was 64 (IQR: 50 -73) with 57.4% older than &gt;65 years. ECOG performance status of &lt;2 was seen in 77.2% of cases; elevated serum LDH in 32.4%; and elevated serum beta-2-microglobulin in 5.6% (n=11/192). Based on previous data, we evaluated and calculated variables that have been suggested as independent negative prognostic factors for overall survival; the proportion of patients with serum albumin &lt;3.5 mg/dL; LMR &lt;2, NLR &gt;4, and PLR &gt;376 was 34.4%, 10.3%, 9.2%, and 4.7% respectively. With a median follow-up of 45 months, the median 5-year overall survival (OS) rate was 72%. The therapy approaches used, response rates and outcomes with these approaches will be presented at the meeting. Results of the univariate and multivariate analysis are summarized in Table 1. We found that age over 65, ECOG performance status, serum albumin level, beta-2-microglobulin level, LDH ratio, LMR, NLR, PLR, and BCL-2 positivity by immunohistochemistry (IHC) were prognostic factors for OS in the univariate analysis. However, in the multivariate analysis, only NLR, serum albumin level and ECOG performance status were independent factor for worse prognosis. Survival rates were significantly shorter in patients with serum albumin &lt;3.5 g/dL (5-year OS of 49% versus 79%, respectively; p&lt;0.0001); NLR &gt;4 (5-year OS of 46% versus 73%, respectively: p=0.0013); and ECOG performance status ≥2 (5-year OS of 51% versus 74%, respectively; p&lt;0.0001) (Figures 1 to 3). Conclusion: In this large cohort of Latin American patients with early stage DLBCL most patients were older than 65, had nodal disease as the primary site, good performance status, with only a third of patients exhibiting elevated LDH. Moreover, we found serum albumin &lt;3.5 g/dL, NLR &gt;4 and ECOG ≥2 as negative prognostic factors for poor survival in early stage DLBCL. We are currently validating our findings in a prospective cohort of Latin American DLBCL patients and to improve clinical decision-making in those deemed at high-risk for early mortality. Figure 1 Figure 1. Disclosures Otero: Astra Zeneca: Current Employment. Oliver: Roche: Other: conference support and fees ; Abbvie: Other: conference support and fees . Castillo: Abbvie: Consultancy, Research Funding; BeiGene: Consultancy, Research Funding; Pharmacyclics: Consultancy, Research Funding; Janssen: Consultancy; Roche: Consultancy; TG Therapeutics: Research Funding.

Urinary Proteomics of Simulated Firefighting Tasks and Its Relation to Fitness Parameters

Firefighting rescues are high-hazard activities accompanied by uncertainty, urgency, and complexity. Knowledge of the metabolic characteristics during firefighting rescues is of great value. The purpose of this study was to explore the firefighting-induced physiological responses in greater depth. The urine samples of ten firefighters were collected before and after the simulated firefighting, and the proteins in urine samples were identified by the liquid chromatography–mass spectroscopy. Blood lactate and heart rate were measured. There were 360 proteins up-regulated and 265 proteins downregulated after this simulated firefighting. Changes in protein expression were significantly related to acute inflammatory responses, immune responses, complement activation, and oxidative stress. Beta-2-microglobulin (r = 0.76, p < 0.05) and von Willebrand factors (r = 0.81, p < 0.01) were positively correlated with heart rate during simulated firefighting, and carbonic anhydrase 1 (r = 0.67, p < 0.05) were positively correlated with blood lactate after simulated firefighting. These results illustrated that Beta-2-microglobulin, von Willebrand, and carbonic anhydrase 1 could be regarded as important indicators to evaluate exercise intensity for firefighters.

ĐẶC ĐIỂM CỦA BỆNH NHÂN ĐA U TỦY CÓ KHUẾCH ĐẠI 1q21 TẠI BỆNH VIỆN TRUYỀN MÁU HUYẾT HỌC

Mục tiêu: Mô tả đặc điểm lâm sàng, xét nghiệm lúc chẩn đoán và đặc điểm di truyền của bệnh nhân đa u tủy có khuếch đại NST 1q. Phương pháp nghiên cứu: Nghiên cứu hồi cứu cắt ngang mô tả hàng loạt ca. Đối tượng: 95 bệnh nhân đa u tủy mới chẩn đoán tại bệnh viện truyền máu huyết học thỏa tiêu chuẩn chọn mẫu trong khoảng thời gian từ 1/2017 đến 12/2020. Kết quả và bàn luận: Tỉ lệ bệnh nhân mang khuếch đại NST 1q là 29,5% (n = 28). Trong số các bất thường về lâm sàng, triệu chứng thiếu máu và đau nhức xương thường gặp nhất (> 70%), ngoài ra còn có triệu chứng u tương bào, sụt cân, sốt, xuất huyết, không có sự khác biệt giữa 2 nhóm có và không có khuếch đại 1q. Tăng calci máu, tăng Beta-2-microglobulin, giai đoạn bệnh muộn, nhiều bất thường di truyền và bất thường thuộc nhóm nguy cơ cao là những đặc điểm khác biệt đáng kể giữa 2 nhóm nghiên cứu (p < 0.05). Kết luận: Có sự khác biệt về các đặc điểm sinh học của nhóm bệnh nhân có khuếch đại NST 1q, cho thấy nhóm BN nay thuộc nhóm nguy cơ cao.

Assessment of the Effects of SARS-Cov-2 Infection on Multiple Organs Using Laboratory Indices

Background: Severe acute respiratory syndrome coronavirus 2 (SARS-Cov-2) is still a severe threaft for human health currently, and the researches about it is a focus topic worldwide. Aim of the study: In this study, we will collect some laboratory results of the patients with coronavirus disease (COVID-19) to assess the function of liver, heart, kidney and even pancreas. Subjects and Methods: Laboratory results of the patients with COVID-19 are collected. The biochemical indices are classified and used to assess the according function of liver, heart, kidney; meantime, and blood glucose is also observed and taken as an index to roughly evaluate pancreas. Results: There were some indices exhibited abnormal. For patient 1, the ratio of albumin and globulin slightly was lower than the down-limit of reference range. For patient 2, alanine aminotransferase (ALT), aspartate aminotransferase (AST), gamma-glutamyltransferase (GGT), creatine kinase (CK), creatinine kinase-MB isoenzyme (CK-MB), lactate dehydrogenase (LDH), alpha-hydroxybutyric dehydrogenase (HBDH), and beta-2 microglobulin (β2-MG) were respectively higher than the according upper limit of the reference range, while prealbumin (PA) was lower than the down limit. For patient 3, GGT, CK, PA were high than normal range. For patient 4, CK, LDH, HBDH were higher than the upper range. Conclusion: Infection of SARS-Cov-2 could cause liver and heart injury, and it is suggested that clinicians and researchers should pay more attention on the prevention, treatment and causative mechanism of such an injury.