OBJECTIVE: Schizophrenia (SZ) and bipolar disorders (BP) are chronic and severe neuropsychiatric diseases. These disorders are tightly related to immune deregulations. In the current study, we intended to replicate the previously reported involvement of the soluble HLA-E isoforms (sHLA-E) in the risk of developing the two conditions along with disease severity in a Tunisian population group.
METHODS: One hundred and twenty-four patients with schizophrenia and 121 with bipolar disorder meeting the DSM-IV criteria along 111 healthy controls were included in this present case-control study. The soluble HLA-E isoforms circulating levels were measured using the ELISA method. The statistical analyses were performed using Kruskal-Wallis and Wilcoxon rank sum tests by R software and GraphPad prism 9.
RESULTS: We found that the sHLA-E circulating levels were significantly higher in BP patients as compared to healthy controls (P<0.0001) and that such increases were mainly observed in patients during an acute phase of their disease (P<0.0001). In SZ patients, while we failed to observe an association with the levels of sHLA-E in the entire SZ sample, we found that high sHLA-E levels characterized stabilized patients in comparison with those during an acute episode (P=0.022). Finally, we did not observe any association between sHLA-E circulating levels and symptoms assessed by the classical clinical scales either in BP or SZ patients.
CONCLUSIONS: Overall, the present findings replicate in a Tunisian population group the previously demonstrated implication of sHLA-E circulating levels in the risk of developing BP or SZ in a French patient cohort. Such replication allows to consider HLA-E as a potent and true inflammatory marker in the context of the two disorders.

BACKGROUND: Natural killer cells (NKs) may be involved in multiple myeloma (MM) progression. The present study elucidated the correlation between NKs and the progression of MM using single-cell binding transcriptome probes to identify NK cell-related biomarkers.
METHODS: Single-cell analysis was performed including cell and subtype annotation, cell communication, and pseudotime analysis. Hallmark pathway enrichment analysis of NKs and NKs-related differentially expressed genes (DEGs) were conducted using Gene Ontology, Kyoto Encyclopedia of Genes and Genomes, and protein-protein interaction (PPI) networks. Then, a risk model was structured based on biomarkers identified through univariate Cox regression analysis and least absolute shrinkage and selection operator regression analysis and subsequently validated. Additionally, correlation of clinical characteristics, gene set enrichment analysis, immune analysis, regulatory network, and drug forecasting were explored.
RESULTS: A total of 13 cell clusters were obtained and annotated, including 8 cell populations that consisted of NKs. Utilizing 123 PPI network node genes, 8 NK-related DEGs were selected to construct a prognostic model. Immune cell infiltration results suggested that 11 immune cells exhibited marked differences in the high and low-risk groups. Finally, the model was used to screen potential drug targets to enhance immunotherapy efficacy.
CONCLUSIONS: A new prognostic model for MM associated with NKs was constructed and validated. This model provides a fresh perspective for predicting patient outcomes, immunotherapeutic response, and candidate drugs.

OBJECTIVE: Onion, particularly onion peel, is a quercetin-rich food with, anti-inflammatory and immunomodulatory effects. However, the effect of onion peel extract (OPE) in humans is unclear. Thus, the present study aimed to investigate whether OPE improves natural killer (NK) cell activity and cytokine concentration in a randomized double-blind placebo-controlled trial.
METHODS: Eighty participants aged 19-64 yrs old with a white blood cell count of 4,000-10,000 cells/µL, symptoms of upper respiratory infection at least once within the previous 12 mon, and perceived stress scale (PSS) over 14 were included. Participants were randomly assigned to take either 1,000 mg/day OPE or a placebo for 8 weeks.
RESULTS: Compliance were 87.4 ± 8.6% and 86.9 ± 79.0% in OPE and placebo groups. Compared to the placebo, OPE supplementation improved \"Hoarseness\" (P = 0.038) of the Wisconsin Upper Respiratory Symptom Survey (WURSS)-21 symptom, and stress scores (P = 0.001; 0.021) of PSS. Supplementation of OPE had no significant effect on NK cell activity and concentrations of cytokines such as interleukin (IL)-2, IL-6, IL-12, IL-1β, interferon-γ, and tumor necrosis factor-α. At baseline, the WURSS-21 symptom and PSS score (P = 0.024; 0.026) were higher in the OPE group than the placebo group. Among participants with higher than median WURSS-21 symptom score, OPE supplementation increased NK cell activity (P = 0.038). Supplementation of OPE had no significant effects on safety measurements and adverse events.
CONCLUSIONS: The present study suggested that OPE supplementation improves NK cell activity in participants with moderate upper respiratory symptoms without any significant adverse effects.
UNASSIGNED: ClinicalTrials.gov Identifier: NCT05666752.

Background Natural killer cells (NK cells) are important mediators of innate immune regulation and literature has shown that they have a role in shaping the adaptive immune system. Objective The present study was undertaken to analyze the NK cell count in systemic lupus erythematosus (SLE) patients as compared to that of controls. Materials and methods Safety of Estrogens in Lupus Erythematosus National Assessment SLE Disease Activity Index (SELENA-SLEDAI) score was assessed in 32 SLE cases. CD3(-) cells were identified as NK cells on flow cytometry, and then their subsets CD56(+) and CD16(+) cells were identified compared to 30 healthy controls. Receiver Operating Characteristic (ROC) curve analysis was performed on NK cells to attempt to determine a cut-off point. Results The CD3(-) NK cells, including the percentages of CD56(+) and CD16(+), were significantly (p<0.001) reduced in SLE patients (12.35%, and 18.7%) as compared to controls (24.67%, and 46.6%). On ROC curve analysis, cut-off values <481/cumm with sensitivity of 86.7% and specificity of 84.4% for CD3(-) NK cells (p<0.001), <23% with 60% sensitivity and 75% specificity for CD56(+) NK cells (p<0.001), and <29% with sensitivity of 70% and specificity of 87.5% for CD16(+) NK cells (p<0.001) were noted. Subsets of NK cells showed no association with the clinicopathological parameters like age, sex, disease activity, anti-nuclear antibodies (ANA), dsDNA, absolute lymphocyte count, and renal involvement. Conclusion NK cells, and their subpopulations of CD56(+) and CD16(+) cells, are decreased in patients with SLE as compared to controls.

UNASSIGNED: Feline chronic gingivostomatitis (FCGS) is a frequent chronic inflammatory condition in the oral cavity with an etiopathogenesis not completely identified. This study aimed to contribute to the knowledge of FCGS by identifying the presence of feline calicivirus (FCV) antigens and natural killer (NK) cells and comparing them.
UNASSIGNED: Forty biopsies from the oral mucosa of cats diagnosed with chronic gingivostomatitis were subjected to immunohistochemical techniques to evaluate cells with FCV antigens and NK cells positive for CD56.
UNASSIGNED: NK cells were identified in all samples, with an average of 725.3 ± 409.1 cells. Regarding FCV, it was identified in 18 out of 30 samples (60%), with a different number of cells with virus in between the analyzed cases. In all cases, the number of cells infected with FCV was lower than the number of NK cells present in the same samples, but there was no statistical association between them.
UNASSIGNED: This preliminary study shows that NK cells are present in gingivostomatitis lesions not exclusively caused by FCV-stimulus, as only 60% of all cases were positive for this virus, but other antigens should be considered in the etiology of FCGS.

Exosomes are released by various cells, including natural killer (NK) cells and transport signaling molecules for the intercellular communication. Hepatocellular carcinoma (HCC), also known as primary liver cancer, is often inoperable and difficult to accurate diagnosis. Notably, the prognosis and underlying mechanisms of HCC are not fully understood. Exosomes-derived NK cells (NK-exos) express unique cytotoxic proteins with a killing ability in tumors and can easily penetrate tumor tissues to improve their targeting ability. NK cell functions, inducing cellular cytotoxicity are modulated by cytokines such as interleukin (IL)-15 and IL-21. However, the mechanisms and effects of cytokines-stimulated NK-exos for the treatment of liver cancer, including HCC, are not well known. In this study, we aimed to investigate the synergistic anti-tumor effects of NK-exos stimulated with IL-15 and IL-21 (NK-exosIL-15/21) in Hep3B cells. Our findings revealed that NK-exosIL-15/21 expressed cytotoxic proteins (perforin and granzyme B) and contained typical exosome markers (CD9 and CD63) within the size range of 100-150 nm. Moreover, we demonstrated that NK-exosIL-15/21 induced the enhancement of cytotoxicity and apoptotic activity in Hep3B cells by activating the specific pro-apoptotic proteins (Bax, cleaved caspase 3, cleaved PARP, perforin, and granzyme B) and inhibiting the anti-apoptotic protein (Bcl-2). In summary, our results suggest that NK-exosIL-15/21 regulate strong anti-tumor effects of HCC cells, by increasing the cytotoxicity and apoptosis through the activation of specific cytotoxic molecules.

Oral squamous cell carcinoma (OSCC) accounts for approximately 90% of oral malignancies and has a 5-year mortality rate close to 50%. A consistent part (70%) of all oral cancers is diagnosed at an advanced stage since available screening techniques are ineffective. Therefore, it would be urgent to improve them. The diagnostic gold standard is tissue biopsy with histological and immunohistochemical assessment. This method presents some limitations. Biopsy is invasive and the histopathological evaluation is semi-quantitative, and the absolute abundance of the target cannot be reliably determined. In addition, tissue is highly processed and may lead to loss of information of the natural state. The search for classical and new clinical biomarkers on fragments of tissue/cells collected with a cytobrush is a highly hopeful technique for early detection and diagnosis of OSCC, because of its non-invasive sampling and easy collection method.
Here we analyzed cytobrush biopsies samples collected from the oral cavity of 15 patients with already diagnosed OSCC by applying an innovative high-sensitivity ELISA technique, in order to verify if this approach may provide useful information for detection, diagnosis, and prognosis of OSCC. To this end, we selected six biomarkers, already used in clinical practice for the diagnosis of OSCC (EGFR, Ki67, p53) or selected based on recent scientific and clinical data which indicate their presence or over-expression in cells undergoing transformation and their role as possible molecular targets in immunecheckpoints blockade therapies (PD-L1, HLA-E, B7-H6).
The selected tumor biomarkers were highly expressed in the tumor core, while were virtually negative in healthy tissue collected from the same patients. These differences were highly statistically significant and consistent with those obtained using the gold standard test clearly indicating that the proposed approach, i.e. analysis of biomarkers by a custom ELISA technique, is strongly reliable.
These preliminary data suggest that this non-invasive rapid phenotyping technique could be useful as a screening tool for phenotyping oral lesions and support clinical practice by precise indications on the characteristics of the lesion, also with a view to the application of new anti-tumor treatments, such as immunotherapy, aimed at OSCC patients.

We investigated the effects of sevoflurane exposure on the expression of matrix metalloproteinase (MMP), expression and ablation of natural killer group 2, member D (NKG2D) ligands (UL16-binding proteins 1-3 and major histocompatibility complex class I chain-related molecules A/B), and natural killer (NK) cell-mediated cytotoxicity in breast cancer cells.
Three human breast cancer cell lines (MCF-7, MDA-MB-453, and HCC-70) were incubated with 0 (control), 600 (S6), or 1200 μM (S12) sevoflurane for 4 h. The gene expression of NKG2D ligands and their protein expression on cancer cell surfaces were measured using multiplex polymerase chain reaction (PCR) and flow cytometry, respectively. Protein expression of MMP-1 and -2 and the concentration of soluble NKG2D ligands were analyzed using western blotting and enzyme-linked immunosorbent assays, respectively.
Sevoflurane downregulated the mRNA and protein expression of the NKG2D ligand in a dose-dependent manner in MCF-7, MDA-MB-453, and HCC-70 cells but did not affect the expression of MMP-1 or -2 or the concentration of soluble NKG2D ligands in the MCF-7, MDA-MB-453, and HCC-70 cells. Sevoflurane attenuated NK cell-mediated cancer cell lysis in a dose-dependent manner in MCF-7, MDA-MB-453, and HCC-70 cells (P = 0.040, P = 0.040, and P = 0.040, respectively).
Our results demonstrate that sevoflurane exposure attenuates NK cell-mediated cytotoxicity in breast cancer cells in a dose-dependent manner. This could be attributed to a sevoflurane-induced decrease in the transcription of NKG2D ligands rather than sevoflurane-induced changes in MMP expression and their proteolytic activity.

Since the first studies, the mouse models have provided crucial support for the most important discoveries on NK cells, on their development, function, and circulation within normal and tumor tissues. Murine tumor models were initially set to study murine NK cells, then, ever more sophisticated human-in-mice models have been developed to investigate the behavior of human NK cells and minimize the interferences from the murine environment. This review presents an overview of the models that have been used along time to study NK cells, focusing on the most popular NOG and NSG models, which work as recipients for the preparation of human-in-mice tumor models, the study of transferred human NK cells, and the evaluation of various enhancers of human NK cell function, including cytokines and chimeric molecules. Finally, an overview of the next generation humanized mice is also provided along with a discussion on how traditional and innovative in-vivo and in-vitro approaches could be integrated to optimize effective pre-clinical studies.

Molecular scale nanopatterns of bioactive molecules have been used to study the effect of transmembrane receptor arrangement on a variety of cell types, including immune cells and their immune response in particular. However, state-of-the-art fabrication approaches have thus far enabled the production of patterns with control over one receptor type only. Herein, we describe a protocol to fabricate arrays for the molecular scale control of the segregation between activating and inhibitory receptors in NK cells. We used this platform to study how ligand segregation regulates NK cell inhibitory signaling and function. The arrays are based on patterns of nanodots of two metals, selectively functionalized with activating and inhibitory ligands. Due to the versatility of our functionalization approach, this protocol can be applied to configurate virtually any combination of extracellular ligands into controlled multifunctional arrays.