PD-L1 Overexpression in the Lungs of Subjects Who Died from COVID-19: Are We on the Way to Understanding the Immune System Exhaustion Induced by SARS-CoV-2?

Knowledge of the pathogenic mechanisms of severe acute respiratory syndrome-associated coronavirus 2 (SARS-CoV-2) is certainly a priority for the scientific community. Two main elements are involved in the biology of the most severe forms of coronavirus disease 2019 (COVID-19): the direct cytopathic effect of the virus against the host tissues, and a dysfunction of the immune system, characterized by the exhaustion of T lymphocytes. The exhaustion of T cells in COVID-19 is poorly understand, but some data could suggest a possible role of PD-1/PD-L1 axis. The aim of this study was to evaluate the possible role of PD-L1 expression in the pulmonary tissue in subjects affected by COVID-19. The presence of SARS-CoV-2 in the pulmonary tissue, and its exact location, was indagated by in situ hybridization; the expression of PD-L1 and CD8 in the same tissue was indagated by immunohistochemistry. Overall, PD-L1 resulted diffusely expressed in 70% of the cases, and an intense expression was observed in 43.5% of cases. Diffuse and intense presence of SARS-CoV-2 by in situ hybridization significantly correlated with an intense PD-L1 expression, and with expression of PD-L1 by pneumocytes. PD-L1 is overexpressed in the pulmonary tissue of subjects died from COVID-19, and mainly in subjects with a high viral load. These data suggest a possible role of PD-L1 in the immune system exhaustion at the basis of the severe forms of the disease.

Brazilian Adaptation and Validation of the Religious and Spiritual Struggles (RSS) Scale—Extended and Short Version

Studies highlight that although the spiritual/religious dimension is commonly used as a resource for coping with stress and suffering, sometimes this dimension can also be a locus of struggles. The Religious and Spiritual Struggles (RSS) Scale, composed of 26 items (extended version), is an instrument that assesses the presence of six types of spiritual struggles, categorized into three major categories: Intrapersonal, Interpersonal, and Supernatural. More recently, studies have presented a leaner version, with 14 items (short version), also suitable for evaluating religious/spiritual struggles. This study aimed to validate the RSS Scale, in its two versions, in the Brazilian cultural context and was able to attest to its quality and reliability, albeit with small variations. The Brazilian extended version presented six components (similar to the original version) explaining 68.48% of variance and 0.907 of KMO. Two items were withdrawn due to low loadings, but the psychometric qualities of both versions in two different samples (one wide sample and one with LGBTQIA+ individuals) were maintained. The reliability (Cronbach’s alpha) for each one of the six components ranged from 0.78 to 0.88 (24 items) and from 0.73 to 0.83 (14 items). The TLI and CFI were, respectively, 0.934 and 0.945 (24 items) and 0.982 and 0.988 for the short version (14 items). Furthermore, convergent validity tests indicated correlations with the Satisfaction with Life Scale. The RSS Scale (the Brazilian versions with 24 and 14 items), demonstrated reliability in the studied sample and can be applied in clinical and research contexts.

Variable levels of spike and ORF1ab RNA in post-mortem lung samples of SARS-CoV-2-positive subjects: comparison between ISH and RT-PCR.

Post-mortem examination plays a pivotal role in understanding the pathobiology of the SARS-CoV-2; thus, the optimization of virus detection on the post-mortem formalin-fixed paraffin-embedded (FFPE) tissue is needed. Different techniques are available for the identification of the SARS-CoV-2, including reverse transcription polymerase chain reaction (RT-PCR), immunohistochemistry (IHC), in situ hybridization (ISH), and electron microscopy. The main goal of this study is to compare ISH versus RT-PCR to detect SARS-CoV-2 on post-mortem lung samples of positive deceased subjects. A total of 27 samples were analyzed by RT-PCR targeting different viral RNA sequences of SARS-CoV-2, including envelope (E), nucleocapsid (N), spike (S), and open reading frame (ORF1ab) genes and ISH targeting S and Orf1ab. All 27 cases showed the N gene amplification, 22 out of 27 the E gene amplification, 26 out of 27 the S gene amplification, and only 6 the ORF1ab gene amplification. The S ISH was positive only in 12 out of 26 cases positive by RT-PCR. The S ISH positive cases with strong and diffuse staining showed a correlation with low values of the number of the amplification cycles by S RT-PCR suggesting that ISH is a sensitive assay mainly in cases carrying high levels of S RNA. In conclusion, our findings demonstrated that ISH assay has lower sensitivity to detect SARS-CoV-2 in FFPE compared to RT-PCR; however, it is able to localize the virus in the cellular context since it preserves the morphology.

Glycated ACE2 receptor in diabetes: open door for SARS-COV-2 entry in cardiomyocyte.

RATIONALE: About 50% of hospitalized coronavirus disease 2019 (COVID-19) patients with diabetes mellitus (DM) developed myocardial damage. The mechanisms of direct SARS-CoV-2 cardiomyocyte infection include viral invasion via ACE2-Spike glycoprotein-binding. In DM patients, the impact of glycation of ACE2 on cardiomyocyte invasion by SARS-CoV-2 can be of high importance. OBJECTIVE: To evaluate the presence of SARS-CoV-2 in cardiomyocytes from heart autopsy of DM cases compared to Non-DM; to investigate the role of DM in SARS-COV-2 entry in cardiomyocytes. METHODS AND RESULTS: We evaluated consecutive autopsy cases, deceased for COVID-19, from Italy between Apr 30, 2020 and Jan 18, 2021. We evaluated SARS-CoV-2 in cardiomyocytes, expression of ACE2 (total and glycosylated form), and transmembrane protease serine protease-2 (TMPRSS2) protein. In order to study the role of diabetes on cardiomyocyte alterations, independently of COVID-19, we investigated ACE2, glycosylated ACE2, and TMPRSS2 proteins in cardiomyocytes from DM and Non-DM explanted-hearts. Finally, to investigate the effects of DM on ACE2 protein modification, an in vitro glycation study of recombinant human ACE2 (hACE2) was performed to evaluate the effects on binding to SARS-CoV-2 Spike protein. The authors included cardiac tissue from 97 autopsies. DM was diagnosed in 37 patients (38%). Fourth-seven out of 97 autopsies (48%) had SARS-CoV-2 RNA in cardiomyocytes. Thirty out of 37 DM autopsy cases (81%) and 17 out of 60 Non-DM autopsy cases (28%) had SARS-CoV-2 RNA in cardiomyocytes. Total ACE2, glycosylated ACE2, and TMPRSS2 protein expressions were higher in cardiomyocytes from autopsied and explanted hearts of DM than Non-DM. In vitro exposure of monomeric hACE2 to 120 mM glucose for 12 days led to non-enzymatic glycation of four lysine residues in the neck domain affecting the protein oligomerization. CONCLUSIONS: The upregulation of ACE2 expression (total and glycosylated forms) in DM cardiomyocytes, along with non-enzymatic glycation, could increase the susceptibility to COVID-19 infection in DM patients by favouring the cellular entry of SARS-CoV2.

How Fear of COVID-19 Can Affect Treatment Choices for Anaplastic Large Cell Lymphomas ALK+ Therapy: A Case Report.

BACKGROUND: The t (2; 5) chromosomal rearrangement of the ALK gene with nucleophosmin 1 gene (NPM1), resulting in an NPM1-ALK fusion, was first demonstrated in 1994 in anaplastic large cell lymphoma, (ALCL), a T-cell lymphoma responsive to cyclophosphamide, abriblastine, vincristine and prednisone in approximately 80% of cases; refractory cases usually respond favorably to brentuximab vedotin. These treatments are regarded as a bridge to allogeneic hematopoietic stem cell transplantation (allo-SCT). Nowadays, transplant procedures and the monitoring of chemotherapy patients proceed very slowly because the SARS-CoV-2 pandemic has heavily clogged the hospitals in all countries. RESULTS: A 40-year-old Caucasian woman was first seen at our clinical center in June 2020. She had ALCL ALK+, a history of failure to two previous therapeutic lines and was in complete remission after 12 courses of brentuximab, still pending allo-SCT after two failed donor selections. Facing a new therapeutic failure, we requested and obtained authorization from the Italian drug regulatory agency to administer 250 mg of crizotinib twice a day, a drug incomprehensibly not registered for ALCL ALK +. CONCLUSIONS: The response to crizotinib was optimal since no adverse event occurred, and CT-PET scans persisted negative; this drug has proved to be a valid bridge to allo-SCT.

Rescue Therapy of Refractory Diffuse Large B-Cell Lymphomas BCL2 with Venetoclax: Case Report.

Eleven years ago, a 64-year-old Caucasian man had LNH Follicular 3a, IV A stage, FLIPI 2 as a prognostic index of follicular lymphoma. He received 8 cycles of RCHOP followed by rituximab maintenance, with complete remission. Due to a systemic recurrence, a new treatment schedule (RCOMP, 6 cycles) was introduced with partial remission persisting during a long-term maintenance treatment with rituximab. Three years ago, LNH Follicular 3a progressed into GC type diffuse large B-cell lymphomas (DLBCL); 6 cycles of rituximab and bendamustine were followed by R-ICE and R OXALI DHAP treatments without beneficial effect. Due to the worse general condition (ECOG 3-4), the patient was treated with pixantrone (6 cycles) until July 10, 2019, with a partial response. On Jan 13, 2020, an extreme compassioned treatment with venetoclax alone was started; this drug was well tolerated and provided a satisfactory clinical and laboratory improvement. In June 2020, however, he developed bone marrow toxicity and septic fever. Nasal and pharyngeal secretions were SARS-CoV-2 RNA negative. Blood cultures for mycotic agents and Gram-positive, Gram-negative, and anaerobic bacteria were negative, but few days later, the patients died of sepsis due to unidentified agents. The use of venetoclax as a single drug to treat DLBCL BCL2 patients deserves further investigation.

Interventional cytopathology in the COVID-19 era.

INTRODUCTION: The dramatic spread of COVID-19 has raised many questions about cytological procedures performed in and out of the laboratories all over the world. METHODS: We report a heterogeneous series of fine needle aspirations performed during the period of phase 1 of the lockdown for the COVID-19 pandemic to describe our experience and measures taken during this period. RESULTS: A total of 48 fine needle aspirations (ultrasound, computed tomography and endoscopic ultrasound guided) were processed and reported. CONCLUSIONS: Pre-existing procedures have been modified to allow healthcare professionals to work safely ensuring patients the necessary assistance with samples suitable for cellularity, fixation and staining for an accurate cytological diagnosis.