Alterations in Alzheimer's disease microglia transcriptome might be involved in bone pathophysiology.

Aging is a major risk factor for multiple chronic disorders in the elderly population, including Alzheimer's disease (AD) and Osteoporosis. AD is a progressive neurodegenerative disease characterized by memory loss. In addition to dementia, several studies have shown that AD patients experience an increased rate of musculoskeletal co-morbidities, such as osteoporosis. Since tissue-specific macrophages contribute to both diseases, this study analyzed the microglia transcriptome of AD mice to determine a common gene signature involved in osteoclast biology. After comparing differentially regulated genes from GEO data sets (GSE93824 and GSE212277), there were 35 common upregulated genes and 89 common downregulated genes. Of these common genes, seven genes are known to play an important role in bone homeostasis. CSF1, SPP1, FAM20C, and Cst7 were upregulated and are associated with osteoclastogenesis and inflammation. Among the downregulated genes, LILRA6, MMP9, and COL18A1 are involved in bone formation and osteoclast regulation. We further validated some of these genes (CSF1, Cst7, and SPP1) in the cortex and the bone of AD mice models. The dysregulation of these microglial genes in AD might provide insights into the co-occurrence of AD and osteoporosis and offer potential therapeutic targets to combat disease progression.

Ethno-demographic disparities in humoral responses to the COVID-19 vaccine among healthcare workers.

The COVID-19 pandemic had a profound impact on global health, but rapid vaccine administration resulted in a significant decline in morbidity and mortality rates worldwide. In this study, we sought to explore the temporal changes in the humoral immune response against SARS-CoV-2 healthcare workers (HCWs) in Augusta, GA, USA, and investigate any potential associations with ethno-demographic features. Specifically, we aimed to compare the naturally infected individuals with naïve individuals to understand the immune response dynamics after SARS-CoV-2 vaccination. A total of 290 HCWs were included and assessed prospectively in this study. COVID status was determined using a saliva-based COVID assay. Neutralizing antibody (NAb) levels were quantified using a chemiluminescent immunoassay system, and IgG levels were measured using an enzyme-linked immunosorbent assay method. We examined the changes in antibody levels among participants using different statistical tests including logistic regression and multiple correspondence analysis. Our findings revealed a significant decline in NAb and IgG levels at 8-12 months postvaccination. Furthermore, a multivariable analysis indicated that this decline was more pronounced in White HCWs (odds ratio [OR] = 2.1, 95% confidence interval [CI] = 1.07-4.08, p = 0.02) and IgG (OR = 2.07, 95% CI = 1.04-4.11, p = 0.03) among the whole cohort. Booster doses significantly increased IgG and NAb levels, while a decline in antibody levels was observed in participants without booster doses at 12 months postvaccination. Our results highlight the importance of understanding the dynamics of immune response and the potential influence of demographic factors on waning immunity to SARS-CoV-2. In addition, our findings emphasize the value of booster doses to ensure durable immunity.

A Comparative Analysis of the Altered Levels of Human Seminal Plasma Constituents as Contributing Factors in Different Types of Male Infertility.

(1) Background: The relationships between the biochemical and immunological components in seminal plasma and their physiological effects on male reproductive system have been underreported. In this study, we evaluated the potential of several seminal plasma biochemical and immunological markers in the pathophysiological developments of the infertile male patients. The study was designed to identify and assess different markers that may be associated with semen functions in different types of male infertility. (2) Methods: A total of 50 infertile male patients who underwent checkup for fertility assessment and 50 fertile controls were included in this study. The complete medical history of each recruited participant was reviewed. The infertile sub-groups (non-obstructive azoospermia (NOA), asthenozoospermia (AS), normozoospermic infertile (NI), and oligozoospermia (OZ)) were characterized based on sperm motility and concentration, while NI patients were included after a thorough check up of their female partners as well. We investigated each sample for 21 different analytes, enzymes, trace elements, and immunological markers to find crucial markers posing as contributing factors to a specific type of male infertility. (3) Results: The levels of 15 out of 21 markers, assayed from the seminal plasma of infertile males, were significantly altered in comparison to fertile controls (p < 0.05). For the first time, microprotein levels were also analyzed. The presence of monocytes, lymphocytes, and granulocytes was limited to semen from NOA patients, while a significant increase in the level of platelets was observed in AS. Hierarchical clustering and ROC-AUC analysis identified the three most significant markers (zinc, LDH, and TG) for the healthy control group and asthenozoospermic group (AUC, of 0.92 and 0.81, respectively). (4) Conclusions: The altered levels of biochemical and immunological markers in seminal plasma might be associated with the different male infertility profiles and could be required for the sperm metabolism and maintenance. However, a larger sample size and follow up analysis is required for establishing the hypothesized panel of markers as biomarkers at clinical stage.

Identification and Clinical Validation of a Novel 4 Gene-Signature with Prognostic Utility in Colorectal Cancer.

Colorectal cancer (CRC) is a high burden disease with several genes involved in tumor progression. The aim of the present study was to identify, generate and clinically validate a novel gene signature to improve prediction of overall survival (OS) to effectively manage colorectal cancer. We explored The Cancer Genome Atlas (TCGA), COAD and READ datasets (597 samples) from The Protein Atlas (TPA) database to extract a total of 595 candidate genes. In parallel, we identified 29 genes with perturbations in > 6 cancers which are also affected in CRC. These genes were entered in cBioportal to generate a 17 gene panel with highest perturbations. For clinical validation, this gene panel was tested on the FFPE tissues of colorectal cancer patients (88 patients) using Nanostring analysis. Using multivariate analysis, a high prognostic score (composite 4 gene signature-DPP7/2, YWHAB, MCM4 and FBXO46) was found to be a significant predictor of poor prognosis in CRC patients (HR: 3.42, 95% CI: 1.71-7.94, p < 0.001 *) along with stage (HR: 4.56, 95% CI: 1.35-19.15, p = 0.01 *). The Kaplan-Meier analysis also segregated patients on the basis of prognostic score (log-rank test, p = 0.001 *). The external validation using GEO dataset (GSE38832, 122 patients) corroborated the prognostic score (HR: 2.7, 95% CI: 1.99-3.73, p < 0.001 *). Additionally, higher score was able to differentiate stage II and III patients (130 patients) on the basis of OS (HR: 2.5, 95% CI: 1.78-3.63, p < 0.001 *). Overall, our results identify a novel 4 gene prognostic signature that has clinical utility in colorectal cancer.

Neurological Complications of COVID-19: Unraveling the Pathophysiological Underpinnings and Therapeutic Implications.

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), the causative agent of coronavirus disease (COVID-19), induced a global pandemic with a diverse array of clinical manifestations. While the acute phase of the pandemic may be waning, the intricacies of COVID-19's impact on neurological health remain a crucial area of investigation. Early recognition of the spectrum of COVID-19 symptoms, ranging from mild fever and cough to life-threatening respiratory distress and multi-organ failure, underscored the significance of neurological complications, including anosmia, seizures, stroke, disorientation, encephalopathy, and paralysis. Notably, patients requiring intensive care unit (ICU) admission due to neurological challenges or due to them exhibiting neurological abnormalities in the ICU have shown increased mortality rates. COVID-19 can lead to a range of neurological complications such as anosmia, stroke, paralysis, cranial nerve deficits, encephalopathy, delirium, meningitis, seizures, etc., in affected patients. This review elucidates the burgeoning landscape of neurological sequelae associated with SARS-CoV-2 infection and explores the underlying neurobiological mechanisms driving these diverse manifestations. A meticulous examination of potential neuroinvasion routes by SARS-CoV-2 underscores the intricate interplay between the virus and the nervous system. Moreover, we dissect the diverse neurological manifestations emphasizing the necessity of a multifaceted approach to understanding the disease's neurological footprint. In addition to elucidating the pathophysiological underpinnings, this review surveys current therapeutic modalities and delineates prospective avenues for neuro-COVID research. By integrating epidemiological, clinical, and diagnostic parameters, we endeavor to foster a comprehensive analysis of the nexus between COVID-19 and neurological health, thereby laying the groundwork for targeted therapeutic interventions and long-term management strategies.

Incorporating Novel Technologies in Precision Oncology for Colorectal Cancer: Advancing Personalized Medicine.

Colorectal cancer (CRC) is one of the most heterogeneous and deadly diseases, with a global incidence of 1.5 million cases per year. Genomics has revolutionized the clinical management of CRC by enabling comprehensive molecular profiling of cancer. However, a deeper understanding of the molecular factors is needed to identify new prognostic and predictive markers that can assist in designing more effective therapeutic regimens for the improved management of CRC. Recent breakthroughs in single-cell analysis have identified new cell subtypes that play a critical role in tumor progression and could serve as potential therapeutic targets. Spatial analysis of the transcriptome and proteome holds the key to unlocking pathogenic cellular interactions, while liquid biopsy profiling of molecular variables from serum holds great potential for monitoring therapy resistance. Furthermore, gene expression signatures from various pathways have emerged as promising prognostic indicators in colorectal cancer and have the potential to enhance the development of equitable medicine. The advancement of these technologies for identifying new markers, particularly in the domain of predictive and personalized medicine, has the potential to improve the management of patients with CRC. Further investigations utilizing similar methods could uncover molecular subtypes specific to emerging therapies, potentially strengthening the development of personalized medicine for CRC patients.

The Role of Aryl Hydrocarbon Receptor in Bone Biology.

Aryl hydrocarbon receptor (AhR), a ligand-activated transcription factor, is crucial in maintaining the skeletal system. Our study focuses on encapsulating the role of AhR in bone biology and identifying novel signaling pathways in musculoskeletal pathologies using the GEO dataset. The GEO2R analysis identified 8 genes (CYP1C1, SULT6B1, CYB5A, EDN1, CXCR4B, CTGFA, TIPARP, and CXXC5A) involved in the AhR pathway, which play a pivotal role in bone remodeling. The AhR knockout in hematopoietic stem cells showed alteration in several novel bone-related transcriptomes (eg, Defb14, ZNF 51, and Chrm5). Gene Ontology Enrichment Analysis demonstrated 54 different biological processes associated with bone homeostasis. Mainly, these processes include bone morphogenesis, bone development, bone trabeculae formation, bone resorption, bone maturation, bone mineralization, and bone marrow development. Employing Functional Annotation and Clustering through DAVID, we further uncovered the involvement of the xenobiotic metabolic process, p450 pathway, oxidation-reduction, and nitric oxide biosynthesis process in the AhR signaling pathway. The conflicting evidence of current research of AhR signaling on bone (positive and negative effects) homeostasis may be due to variations in ligand binding affinity, binding sites, half-life, chemical structure, and other unknown factors. In summary, our study provides a comprehensive understanding of the underlying mechanisms of the AhR pathway in bone biology.

Single episode of moderate to severe traumatic brain injury leads to chronic neurological deficits and Alzheimer's-like pathological dementia.

Traumatic brain injury (TBI) is one of the foremost causes of disability and mortality globally. While the scientific and medical emphasis is to save lives and avoid disability during acute period of injury, a severe health problem can manifest years after injury. For instance, TBI increases the risk of cognitive impairment in the elderly. Remote TBI history was reported to be a cause of the accelerated clinical trajectory of Alzheimer's disease-related dementia (ADRD) resulting in earlier onset of cognitive impairment and increased AD-associated pathological markers like greater amyloid deposition and cortical thinning. It is not well understood whether a single TBI event may increase the risk of dementia. Moreover, the cellular signaling pathways remain elusive for the chronic effects of TBI on cognition. We have hypothesized that a single TBI induces sustained neuroinflammation and disrupts cellular communication in a way that results later in ADRD pathology. To test this, we induced TBI in young adult CD1 mice and assessed the behavioral outcomes after 11 months followed by pathological, histological, transcriptomic, and MRI assessment. On MRI scans, these mice showed significant loss of tissue, reduced CBF, and higher white matter injury compared to sham mice. We found these brains showed progressive atrophy, markers of ADRD, sustained astrogliosis, loss of neuronal plasticity, and growth factors even after 1-year post-TBI. Because of progressive neurodegeneration, these mice had motor deficits, showed cognitive impairments, and wandered randomly in open field. We, therefore, conclude that progressive pathology after adulthood TBI leads to neurodegenerative conditions such as ADRD and impairs neuronal functions.

Immune Factors Drive Expression of SARS-CoV-2 Receptor Genes Amid Sexual Disparity.

The emergence of COVID-19 has led to significant morbidity and mortality, with around seven million deaths worldwide as of February 2023. There are several risk factors such as age and sex that are associated with the development of severe symptoms due to COVID-19. There have been limited studies that have explored the role of sex differences in SARS-CoV-2 infection. As a result, there is an urgent need to identify molecular features associated with sex and COVID-19 pathogenesis to develop more effective interventions to combat the ongoing pandemic. To address this gap, we explored sex-specific molecular factors in both mouse and human datasets. The host immune targets such as TLR7, IRF7, IRF5, and IL6, which are involved in the immune response against viral infections, and the sex-specific targets such as AR and ESSR were taken to investigate any possible link with the SARS-CoV-2 host receptors ACE2 and TMPRSS2. For the mouse analysis, a single-cell RNA sequencing dataset was used, while bulk RNA-Seq datasets were used to analyze the human clinical data. Additional databases such as the Database of Transcription Start Sites (DBTS), STRING-DB, and the Swiss Regulon Portal were used for further analysis. We identified a 6-gene signature that showed differential expression in males and females. Additionally, this gene signature showed potential prognostic utility by differentiating ICU patients from non-ICU patients due to COVID-19. Our study highlights the importance of assessing sex differences in SARS-CoV-2 infection, which can assist in the optimal treatment and better vaccination strategies.

TIMP-1 Dependent Modulation of Metabolic Profiles Impacts Chemoresistance in NSCLC.

The development of chemoresistance remains a significant barrier to treating NSCLC. Alteration of cancer cell metabolism is an important mechanism for chemoresistance. This study explored the role of aberrant metabolism in TIMP-1-mediated chemoresistance. Bioinformatics analysis identified an association of high TIMP-1 with altered energy metabolism. We have defined the role of depolarized mitochondria through a reduction in lactate secretion, higher ROS levels in TIMP-1 KD cells and reduced GSH levels. TIMP-1 modulates the metabolic profile via acetylation of mitochondrial STAT3 and its interaction with CD44. Intriguingly, monomers of acetylated STAT3 were critical for altered metabolism, whereas STAT3 dimers abrogated this function. Further, the mitochondrial metabolic profile was also altered in a cisplatin-resistant clone of A549 cells. We also correlated the immunoexpression of CD44, STAT3 and TIMP-1 in patient samples. This study provided evidence that TIMP-1 alters the metabolic profile by modulating mitochondrial metabolism via the CD44-STAT3 axis through its effects on STAT3 acetylation. It also lent further support to the critical role of TIMP-1 in chemoresistance. Interrogation of the TCGA-LUAD dataset revealed perturbations in the critical modulator that can alter metabolic states in cancer cells. Higher expression of a five-gene signature, including TIMP-1, correlated with immunosuppressive cells and was found to be associated with overall survival. This study identified several metabolic mechanisms that could influence therapeutic options and prognosis in NSCLC patients.

Clinical validation of a multiplex PCR-based detection assay using saliva or nasopharyngeal samples for SARS-Cov-2, influenza A and B.

The COVID-19 pandemic has resulted in significant diversion of human and material resources to COVID-19 diagnostics, to the extent that influenza viruses and co-infection in COVID-19 patients remains undocumented and pose serious public-health consequences. We optimized and validated a highly sensitive RT-PCR based multiplex-assay for the detection of SARS-CoV-2, influenza A and B viruses in a single-test. This study evaluated clinical specimens (n = 1411), 1019 saliva and 392 nasopharyngeal swab (NPS), tested using two-assays: FDA-EUA approved SARS-CoV-2 assay that targets N and ORF1ab gene, and the PKamp-RT-PCR based assay that targets SARS-CoV-2, influenza viruses A and B. Of the 1019 saliva samples, 17.0% (174/1019) tested positive for SARS-CoV-2 using either assay. The detection rate for SARS-CoV-2 was higher with the multiplex assay compared to SARS-specific assay [91.9% (160/174) vs. 87.9% (153/174)], respectively. Of the 392 NPS samples, 10.4% (41/392) tested positive for SARS-CoV-2 using either assay. The detection rate for SARS-CoV-2 was higher with the multiplex assay compared to SARS-specific assay [97.5% (40/41) vs. 92.1% (39/41)], respectively. This study presents clinical validation of a multiplex-PCR assay for testing SARS-CoV-2, influenza A and B viruses, using NPS and saliva samples, and demonstrates the feasibility of implementing the assay without disrupting the existing laboratory workflow.

Clinical and molecular assessment of an onco-immune signature with prognostic significance in patients with colorectal cancer.

Understanding the complex tumor microenvironment is key to the development of personalized therapies for the treatment of cancer including colorectal cancer (CRC). In the past decade, significant advances in the field of immunotherapy have changed the paradigm of cancer treatment. Despite significant improvements, tumor heterogeneity and lack of appropriate classification tools for CRC have prevented accurate risk stratification and identification of a wider patient population that may potentially benefit from targeted therapies. To identify novel signatures for accurate prognostication of CRC, we quantified gene expression of 12 immune-related genes using a medium-throughput NanoString quantification platform in 93 CRC patients. Multivariate prognostic analysis identified a combined four-gene prognostic signature (TGFB1, PTK2, RORC, and SOCS1) (HR: 1.76, 95% CI: 1.05-2.95, *p < 0.02). The survival trend was captured in an independent gene expression data set: GSE17536 (177 patients; HR: 3.31, 95% CI: 1.99-5.55, *p < 0.01) and GSE14333 (226 patients; HR: 2.47, 95% CI: 1.35-4.53, *p < 0.01). Further, gene set enrichment analysis of the TCGA data set associated higher prognostic scores with epithelial-mesenchymal transition (EMT) and inflammatory pathways. Comparatively, a lower prognostic score was correlated with oxidative phosphorylation and MYC and E2F targets. Analysis of immune parameters identified infiltration of T-reg cells, CD8+ T cells, M2 macrophages, and B cells in high-risk patient groups along with upregulation of immune exhaustion genes. This molecular study has identified a novel prognostic gene signature with clinical utility in CRC. Therefore, along with prognostic features, characterization of immune cell infiltrates and immunosuppression provides actionable information that should be considered while employing personalized medicine.

The clinical relevance of gene expression based prognostic signatures in colorectal cancer.

Colorectal cancer (CRC) is one of the most prevalent cancers, with more than one million new cases every year. In the last few decades, several advancements in therapeutic and preventative levels have reduced the mortality rate, but new biomarkers are required for improved prognosis. The alterations at the genetic and epigenetic level have been recognized as major players in tumorigenesis. The products of gene expression in the form of mRNA, microRNA, and long-noncoding RNA, have started to emerge as important regulatory molecules, playing an important role in cancer. Gene-expression based prognostic risk scores, which quantify and compare their expression, have emerged as promising biomarkers with enormous clinical value. These composite multi-gene models in which more than one gene is used to predict prognosis have been shown to be significantly effective in identifying patients with multiple clinico-pathological risks like overall mortality, response to chemotherapy, risk of metastasis, etc. The advent of microarray and advanced sequencing technologies have led to the generation of large datasets like TCGA (The Cancer Genome Atlas) and GEO (Gene Expression Omnibus), which have fueled the search for new biomarkers. Continuous evaluation of these candidate biomarkers in clinical settings is promising to improve the management of CRC. These composite gene signatures provide potential in identifying high-risk patients, which might help clinicians to better manage these patients and design appropriate personalized therapeutic interventions. In this review, we emphasize on composite prognostic scores from diverse resources with clinical utility in CRC.

Gene signatures with therapeutic value: emerging perspective for personalized immunotherapy in renal cancer.

Renal cancer is one of the deadliest urogenital diseases. In recent years, the advent of immunotherapy has led to significant improvement in the management of patients with renal cancer. Although cancer immunotherapy and its combinations had benefited numerous patients, several challenges need to be addressed. Apart from the high costs of treatment, the lack of predictive biomarkers and toxic side-effects have impeded its wider applicability. To address these issues, new biomarkers are required to predict responsiveness and design personalized treatment strategies. Recent advances in the field of single-cell sequencing and multi-dimensional spatial transcriptomics have identified clinically relevant subtypes of renal cancer. Furthermore, there is emerging potential for gene signatures based on immune cells, non-coding RNAs, and pathways such as metabolism and RNA modification. In this review article, we have discussed recent progress in the identification of gene signatures with predictive and prognostic potential in renal cancer.

Prognostic and therapeutic implications of extracellular matrix associated gene signature in renal clear cell carcinoma.

Complex interactions in tumor microenvironment between ECM (extra-cellular matrix) and cancer cell plays a central role in the generation of tumor supportive microenvironment. In this study, the expression of ECM-related genes was explored for prognostic and immunological implication in clear cell renal clear cell carcinoma (ccRCC). Out of 964 ECM genes, higher expression (z-score > 2) of 35 genes showed significant association with overall survival (OS), progression-free survival (PFS) and disease-specific survival (DSS). On comparison to normal tissue, 12 genes (NUDT1, SIGLEC1, LRP1, LOXL2, SERPINE1, PLOD3, ZP3, RARRES2, TGM2, COL3A1, ANXA4, and POSTN) showed elevated expression in kidney tumor (n = 523) compared to normal (n = 100). Further, Cox proportional hazard model was utilized to develop 12 genes ECM signature that showed significant association with overall survival in TCGA dataset (HR = 2.45; 95% CI [1.78-3.38]; p < 0.01). This gene signature was further validated in 3 independent datasets from GEO database. Kaplan-Meier log-rank test significantly associated patients with elevated expression of this gene signature with a higher risk of mortality. Further, differential gene expression analysis using DESeq2 and principal component analysis (PCA) identified genes with the highest fold change forming distinct clusters between ECM-rich high-risk and ECM-poor low-risk patients. Geneset enrichment analysis (GSEA) identified significant perturbations in homeostatic kidney functions in the high-risk group. Further, higher infiltration of immunosuppressive T-reg and M2 macrophages was observed in high-risk group patients. The present study has identified a prognostic signature with associated tumor-promoting immune niche with clinical utility in ccRCC. Further exploration of ECM dynamics and validation of this gene signature can assist in design and application of novel therapeutic approaches.

Immunogenomic Gene Signature of Cell-Death Associated Genes with Prognostic Implications in Lung Cancer.

Lung cancer is one of the leading causes of death worldwide. Cell death pathways such as autophagy, apoptosis, and necrosis can provide useful clinical and immunological insights that can assist in the design of personalized therapeutics. In this study, variations in the expression of genes involved in cell death pathways and resulting infiltration of immune cells were explored in lung adenocarcinoma (The Cancer Genome Atlas: TCGA, lung adenocarcinoma (LUAD), 510 patients). Firstly, genes involved in autophagy (n = 34 genes), apoptosis (n = 66 genes), and necrosis (n = 32 genes) were analyzed to assess the prognostic significance in lung cancer. The significant genes were used to develop the cell death index (CDI) of 21 genes which clustered patients based on high risk (high CDI) and low risk (low CDI). The survival analysis using the Kaplan-Meier curve differentiated patients based on overall survival (40.4 months vs. 76.2 months), progression-free survival (26.2 months vs. 48.6 months), and disease-free survival (62.2 months vs. 158.2 months) (Log-rank test, p < 0.01). Cox proportional hazard model significantly associated patients in high CDI group with a higher risk of mortality (Hazard Ratio: H.R 1.75, 95% CI: 1.28-2.45, p < 0.001). Differential gene expression analysis using principal component analysis (PCA) identified genes with the highest fold change forming distinct clusters. To analyze the immune parameters in two risk groups, cytokines expression (n = 265 genes) analysis revealed the highest association of IL-15RA and IL 15 (> 1.5-fold, p < 0.01) with the high-risk group. The microenvironment cell-population (MCP)-counter algorithm identified the higher infiltration of CD8+ T cells, macrophages, and lower infiltration of neutrophils with the high-risk group. Interestingly, this group also showed a higher expression of immune checkpoint molecules CD-274 (PD-L1), CTLA-4, and T cell exhaustion genes (HAVCR2, TIGIT, LAG3, PDCD1, CXCL13, and LYN) (p < 0.01). Furthermore, functional enrichment analysis identified significant perturbations in immune pathways in the higher risk group. This study highlights the presence of an immunocompromised microenvironment indicated by the higher infiltration of cytotoxic T cells along with the presence of checkpoint molecules and T cell exhaustion genes. These patients at higher risk might be more suitable to benefit from PD-L1 blockade or other checkpoint blockade immunotherapies.

Infection and Immune Memory: Variables in Robust Protection by Vaccines Against SARS-CoV-2.

SARS-CoV-2 is the cause of a recent pandemic that has led to more than 3 million deaths worldwide. Most individuals are asymptomatic or display mild symptoms, which raises an inherent question as to how does the immune response differs from patients manifesting severe disease? During the initial phase of infection, dysregulated effector immune cells such as neutrophils, macrophages, monocytes, megakaryocytes, basophils, eosinophils, erythroid progenitor cells, and Th17 cells can alter the trajectory of an infected patient to severe disease. On the other hand, properly functioning CD4+, CD8+ cells, NK cells, and DCs reduce the disease severity. Detailed understanding of the immune response of convalescent individuals transitioning from the effector phase to the immunogenic memory phase can provide vital clues to understanding essential variables to assess vaccine-induced protection. Although neutralizing antibodies can wane over time, long-lasting B and T memory cells can persist in recovered individuals. The natural immunological memory captures the diverse repertoire of SARS-CoV-2 epitopes after natural infection whereas, currently approved vaccines are based on a single epitope, spike protein. It is essential to understand the nature of the immune response to natural infection to better identify 'correlates of protection' against this disease. This article discusses recent findings regarding immune response against natural infection to SARS-CoV-2 and the nature of immunogenic memory. More precise knowledge of the acute phase of immune response and its transition to immunological memory will contribute to the future design of vaccines and the identification of variables essential to maintain immune protection across diverse populations.

Natural Killer Cells and Dendritic Cells: Expanding Clinical Relevance in the Non-Small Cell Lung Cancer (NSCLC) Tumor Microenvironment.

Non-small cell lung cancer (NSCLC) is a major subtype of lung cancer that accounts for almost 85% of lung cancer cases worldwide. Although recent advances in chemotherapy, radiotherapy, and immunotherapy have helped in the clinical management of these patients, the survival rate in advanced stages remains dismal. Furthermore, there is a critical lack of accurate prognostic and stratification markers for emerging immunotherapies. To harness immune response modalities for therapeutic benefits, a detailed understanding of the immune cells in the complex tumor microenvironment (TME) is required. Among the diverse immune cells, natural killer (NK cells) and dendritic cells (DCs) have generated tremendous interest in the scientific community. NK cells play a critical role in tumor immunosurveillance by directly killing malignant cells. DCs link innate and adaptive immune systems by cross-presenting the antigens to T cells. The presence of an immunosuppressive milieu in tumors can lead to inactivation and poor functioning of NK cells and DCs, which results in an adverse outcome for many cancer patients, including those with NSCLC. Recently, clinical intervention using modified NK cells and DCs have shown encouraging response in advanced NSCLC patients. Herein, we will discuss prognostic and predictive aspects of NK cells and DC cells with an emphasis on NSCLC. Additionally, the discussion will extend to potential strategies that seek to enhance the anti-tumor functionality of NK cells and DCs.

Guideline-Adherent Clinical Validation of a Comprehensive 170-Gene DNA/RNA Panel for Determination of Small Variants, Copy Number Variations, Splice Variants, and Fusions on a Next-Generation Sequencing Platform in the CLIA Setting.

We describe the clinical validation of a targeted DNA and RNA-based next-generation sequencing (NGS) assay at two clinical molecular diagnostic laboratories. This assay employs simultaneous DNA and RNA analysis of all coding exons to detect small variants (single-nucleotide variants, insertions, and deletions) in 148 genes, amplifications in 59 genes, and fusions and splice variants in 55 genes. During independent validations at two sites, 234 individual specimens were tested, including clinical formalin-fixed, paraffin-embedded (FFPE) tumor specimens, reference material, and cell lines. Samples were prepared using the Illumina TruSight Tumor 170 (TST170) kit, sequenced with Illumina sequencers, and the data were analyzed using the TST170 App. At both sites, TST170 had ≥98% success for ≥250× depth for ≥95% of covered positions. Variant calling was accurate and reproducible at allele frequencies ≥5%. Limit of detection studies determined that inputs of ≥50 ng of DNA (with ≥3.3 ng/µl) and ≥50 ng RNA (minimum of 7 copies/ng) were optimal for high analytical sensitivity. The TST170 assay results were highly concordant with prior results using different methods across all variant categories. Optimization of nucleic acid extraction and DNA shearing, and quality control following library preparation is recommended to maximize assay success rates. In summary, we describe the validation of comprehensive and simultaneous DNA and RNA-based NGS testing using TST170 at two clinical sites.

Nanostring-Based Identification of the Gene Expression Profile in Trigger Finger Samples.

Trigger finger is a common yet vastly understudied fibroproliferative hand pathology, severely affecting patients' quality of life. Consistent trauma due to inadequate positioning within the afflicted finger's tendon/pulley system leads to cellular dysregulation and eventual fibrosis. While the genetic characteristics of the fibrotic tissue in the trigger finger have been studied, the pathways that govern the initiation and propagation of fibrosis are still unknown. The complete gene expression profile of the trigger finger has never been explored. Our study has used the Nanostring nCounter gene expression assay to investigate the molecular signaling involved in trigger finger pathogenesis. We collected samples from patients undergoing trigger finger (n = 4) release surgery and compared the gene expression to carpal tunnel tissue (n = 4). Nanostring nCounter analysis identified 165 genes that were differentially regulated; 145 of these genes were upregulated, whereas 20 genes were downregulated. We found that several collagen genes were significantly upregulated, and a regulatory matrix metalloproteinase (MMP), MMP-3, was downregulated. Bioinformatic analysis revealed that several known signaling pathways were dysregulated, such as the TGF-ß1 and Wnt signaling pathways. We also found several novel signaling pathways (e.g., PI3K, MAPK, JAK-STAT, and Notch) differentially regulated in trigger finger. The outcome of our study helps in understanding the molecular signaling pathway involved in the pathogenesis of the trigger finger.