Correlation of Lymphocyte Subpopulations, Clinical Features and Inflammatory Markers during Severe COVID-19 Onset.

BACKGROUND: Dysregulation of the immune response in the course of COVID-19 has been implicated in critical outcomes. Lymphopenia is evident in severe cases and has been associated with worse outcomes since the early phases of the pandemic. In addition, cytokine storm has been associated with excessive lung injury and concomitant respiratory failure. However, it has also been hypothesized that specific lymphocyte subpopulations (CD4 and CD8 T cells, B cells, and NK cells) may serve as prognostic markers for disease severity. The aim of this study was to investigate possible associations of lymphocyte subpopulations alterations with markers of disease severity and outcomes in patients hospitalized with COVID-19. MATERIALS/METHODS: A total of 42 adult hospitalized patients were included in this study, from June to July 2021. Flow-cytometry was used to calculate specific lymphocyte subpopulations on day 1 (admission) and on day 5 of hospitalization (CD45, CD3, CD3CD8, CD3CD4, CD3CD4CD8, CD19, CD16CD56, CD34RA, CD45RO). Markers of disease severity and outcomes included: burden of disease on CT (% of affected lung parenchyma injury), C-reactive protein and interleukin-6 levels. PO2/FiO2 ratio and differences in lymphocytes subsets between two timepoints were also calculated. Logistic and linear regressions were used for the analyses. All analyses were performed using Stata (version 13.1; Stata Corp, College Station, TX, USA). RESULTS: Higher levels of CD16CD56 cells (Natural Killer cells) were associated with higher risk of lung injury (>50% of lung parenchyma). An increase in CD3CD4 and CD4RO cell count difference between day 5 and day 1 resulted in a decrease of CRP difference between these timepoints. On the other hand, CD45RARO difference was associated with an increase in the difference of CRP levels between the two timepoints. No other significant differences were found in the rest of the lymphocyte subpopulations. CONCLUSIONS: Despite a low patient number, this study showed that alterations in lymphocyte subpopulations are associated with COVID-19 severity markers. It was observed that an increase in lymphocytes (CD4 and transiently CD45RARO) resulted in lower CRP levels, perhaps leading to COVID-19 recovery and immune response homeostasis. However, these findings need further evaluation in larger scale trials.

Global profiling of EGFR gene mutation, amplification, regulation and tissue protein expression in unknown primary carcinomas: to target or not to target?

INTRODUCTION: Epidermal growth factor receptor (EGFR) signalling contributes to malignant transformation and survival. We studied molecular predictors of benefit from EGFR-modulating therapies in patients with cancer of unknown primary (CUP). MATERIALS AND METHODS: Tumours from paraffin-embedded biopsies of 50 patients with CUP were stained for EGFR protein by immunohistochemistry. Polymerase chain reaction amplification, single-strand conformational polymorphism and direct sequencing were used to study EGFR intron 1 cytosine-adenosine (CA) repeat length as well as exon 18, 19, 21 activating mutations and amplification. RESULTS: Thirty-seven tumours (74%) expressed EGFR protein but only six (12%) strongly. Regarding intron 1 CA repeat length, we detected five alleles with CA repeat numbers 16-20, allele 16 being the most common (39%). All samples were heterozygous, the commonest genotype consisting of 16/18 dinucleotides (78%). Five samples had three intron 1 alleles and were associated with EGFR overexpression in 40% of cases. There was no evidence of EGFR exon 18, 19, 21 amplification. Two mutations were detected: Exon 21 2508 C > T, a silent nucleotide polymorphism (R836R) and a G > A substitution in sequences flanking exon 19 (IVS19 + 24G > A) resulting in aberrant mRNA splicing. Neither EGFR protein expression nor CA repeat length were prognostic factors for survival. CONCLUSIONS: Our data depict absence of molecular predictors of benefit from EGFR modulation in patients with CUP. Study of its molecular pathophysiology and targeting other molecular pathways may be warranted instead.

DNA content is associated with malignancy of intracranial neoplasms.

OBJECTIVE: Flow cytometry has been applied to analyze the DNA-content distribution of tumors, in order to relate this to clinical and biological parameters of tumor behavior. Herewith, we investigated the value of cell cycle analysis in the characterization of intracranial lesions and its possible prognostic role. METHODS: DNA analysis was performed in tumor samples that were taken during surgery over a five year period. Diagnosed tumors were graded according to the World Health Organization 2007 classification scheme. RESULTS: Fifty-six patients were included in the study. There was a significant difference in G0/G1 phase and S-phase between low-grade and high-grade gliomas. There were 12 (57%) diploid and 9 (43%) aneuploid tumors. All aneuploid tumors were glioblastomas. Patients with G0/G1 value ≤ 69% and S phase value greater than 6% were associated with worse survival. As regards meningiomas, there was a significant difference in G0/G1 phase, S phase and mitoses fraction between benign and both atypical and anaplastic meningiomas. Aneuploidy was observed in the anaplastic tumors and in 2/4 atypical meningiomas. CONCLUSION: The results of the present study, showed that cell cycle analysis could differentiate low from high grade gliomas and benign from atypical/anaplastic meningiomas. Furthermore, a prognostic significance was found in glioma patients. The role of cell cycle analysis in brain tumors thus warrants further investigation.