Cell-mediated Research Articles

IFN‐α treatment may enable discontinuation of TKIs in NK cell‐licensed patients with CML‐CP

AbstractThe magnitude of the natural killer (NK) cell response contributes to the achievement of treatment‐free remission (TFR) in patients with chronic myeloid leukemia (CML) and is regulated by the interaction between killer immunoglobulin‐like receptors (KIRs) on NK cells and human leukocyte antigen (HLA) class I molecules on target cells. The abundant combination between KIR and HLA through genetic polymorphisms determines the functional diversity of NK cells. We previously reported that KIR3DL1‐HLA‐Bw status is associated with achievement of TFR by reflecting NK cell potential. Patients with strong interaction between KIR3DL1/HLA‐Bw were identified as having a higher molecular relapse risk, based on the “missing self” hypothesis which suggests that the lack of cognate ligands for KIRs may induce target cell lysis. However, all the patients with strong interaction between KIR3DL1/HLA‐Bw who received prior IFN‐α therapy achieved TFR (p = 0.007), explained by the “NK cell licensing” concept, whereby NK cells become more functional through the recognition “self” HLA class I molecules by KIRs. NK cell licensing may contribute to the potential efficacy of IFN‐α treatment in patients with CML. We defined high‐risk molecular relapse patients and suggest that KIR3DL1/HLA‐Bw status may help detect patients who could benefit from IFN‐α for maintaining TFR.

High throughput cell mechanotyping of cell response to cytoskeletal modulations using a microfluidic cell deformation system

High throughput cell mechanotyping of cell response to cytoskeletal modulations using a microfluidic cell deformation system

Clonal hematopoiesis impacts frailty in newly diagnosed multiple myeloma patients: a retrospective multicenter analysis

Somatic mutations of hematopoietic cells in the peripheral blood of normal individuals refer to clonal hematopoiesis of indeterminate potential (CHIP), which is associated with a 0.5–1% risk of progression to hematological malignancies and cardiovascular diseases. CHIP has also been reported in Multiple Myeloma (MM) patients, but its biological relevance remains to be elucidated. In this study, high-depth targeted sequencing of peripheral blood from 76 NDMM patients revealed CHIP in 46% of them, with a variant allele frequency (VAF) ranging from ~ 1 to 34%. The most frequently mutated gene was DNMT3A, followed by TET2. More aggressive disease features were observed among CHIP carriers, who also exhibited higher proportion of high-risk stages (ISS and R-ISS 3) compared to controls. Longitudinal analyses at diagnosis and during follow-up showed a slight increase of VAFs (p = 0.058) for epigenetic (DNMT3A, TET2, and ASXL1) and DNA repair genes (TP53; p = 0.0123). A more stable frequency was observed among other genes, suggesting different temporal dynamics of CH clones. Adverse clinical outcomes, in term of overall and progression-free survivals, were observed in CHIP carriers. These patients also exhibited weakened immune T-cells and enhanced frailty, predicting greater toxicity and consequently shorter event-free survival. Finally, correlation analysis identified platelets count as biomarker for higher VAF among CHIP carriers, regardless of the specific variant. Overall, our study highlights specific biological and clinical features, paving the way for the development of tailored strategies for MM patients carrying CHIP.

Chronic inflammation degrades CD4 T cell immunity to prior vaccines in treated HIV infection

To date, our understanding of how HIV infection impacts vaccine-induced cellular immunity is limited. Here, we investigate inflammation, immune activation and antigen-specific T cell responses in HIV-uninfected and antiretroviral-treated HIV-infected people. Our findings highlight lower recall responses of antigen-specific CD4 T cells that correlate with high plasma cytokines levels, T cell hyperactivation and an altered composition of the T subsets enriched with more differentiated cells in the HIV-infected group. Transcriptomic analysis reveals that antigen-specific CD4 T cells of the HIV-infected group have a reduced expression of gene sets previously reported to correlate with vaccine-induced pathogen-specific protective immunity and further identifies a consistent impairment of the IFNα and IFNγ response pathways as mechanism for the functional loss of recall CD4 T cell responses in antiretroviral-treated people. Lastly, in vitro treatment with drugs that reduce inflammation results in higher memory CD4 T cell IFNγ responses. Together, our findings suggest that vaccine-induced cellular immunity may benefit from strategies to counteract inflammation in HIV infection.

A Bioinspired Nanovaccine for Personalized Cancer Immunotherapy.

Poly I:C (pIC) can act on endosomal and cytosolic pathogen recognition receptors to enhance T cell immunity. However, the poor cytosolic delivery of pIC and lack of facile methods for codelivery with antigens limit its efficacy. Inspired by the structure of a virus, we developed a liponanogel (LNG) consisting of a nanogel core and lipid shell to address these challenges. An LNG-based vaccine increases the endosomal membrane permeability in a nanogel core-dependent manner, thus enhancing cytosolic sensing of pIC. LNG induces 44.9-fold stronger CD8+ T cell responses than soluble pIC or Hiltonol adjuvanted vaccines and even induces stronger CD8+ T cell responses than state-of-the-art lipid nanoparticle adjuvanted vaccines. Remarkably, the LNG vaccine regresses 100% TC1 tumors and even regresses 60% aggressive B16F10 tumors upon combination with αPD-L1. Our study provides a safe and effective strategy for enhancing T cell immunity and may inspire new approaches for cancer immunotherapy.

Systematic review and meta-analysis of recombinant herpes zoster vaccine in immunocompromised populations.

Herpes zoster infection is common in immunocompromised individuals. Recently, the Advisory Committee on Immunization Practices recommended immunizing with the recombinant zoster vaccine (RZV). To evaluate the efficacy, immunogenicity and safety of RZV in immunocompromised individuals, such as transplant recipients, cancer patients undergoing chemotherapy, individuals with preexisting autoimmune diseases and HIV-infected patients. From January 1984 to October 2023, a systematic search of PubMed, MEDLINE, EMBASE, Scopus, Web of Science, CINAHL, and Cochrane CENTRAL was performed. Randomized clinical trials (RCT) evaluating RZV compared to placebo in immunocompromised adults were selected. Study characteristics and estimates on the incidence of herpes zoster, immune responses, and safety data were extracted from studies. Estimates were pooled using random-effects meta-analysis. Differences by study-level characteristics were estimated using subgroup meta-analysis and metaregression. Seven RCTs were included. Compared to placebo, RZV reduced the incidence of herpes zoster across all ages by 81% (RR: 0.19, 95%CI: 0.09, 0.44), with moderate heterogeneity across the studies (I2 = 60.49%; τ2 = 0.31; P = 0.07). RZV significantly increased humoral and cellular immunity one month after the last dose. Transplant and past malignancy were associated with lower immunogenicity. RZV was more reactogenic with more local and systemic adverse events. There was no difference in serious adverse events or death between the two arms. This study suggests that RZV reduces the risk of herpes zoster infection in immunocompromised individuals. This vaccine should be routinely offered to immunocompromised individuals, preferably before chemotherapy or treatment.

Diverse priming outcomes under conditions of very rare precursor B cells.

Rare B cells can have special pathogen-recognition features giving them the potential to make outsized contributions to protective immunity. However, rare naive B cells infrequently participate in immune responses. We investigated how germline-targeting vaccine antigen delivery and adjuvant selection affect priming of exceptionally rare BG18-like HIV broadly neutralizing antibody-precursor B cells (~1 in 50 million) in non-human primates. Only escalating dose (ED) priming immunization using the saponin adjuvant SMNP elicited detectable BG18-like cells in germinal centers (GCs). All groups had strong GC responses, but only ED+SMNP and bolus+SMNP induced BG18-like memory B cells in >50% of animals. One group had vaccine-specific GC responses equivalent to ED+SMNP, but BG18-like memory B cells were rarely detected. Following homologous boosting, BG18-like memory B cells were more frequent in a bolus priming group, but had lower somatic hypermutation and affinities. This outcome was inversely associated with post-prime antibody titers, suggesting antibody feedback can significantly influence rare precursor B cell responses.

CCL2-CCR2 Axis Inhibition in Osteosarcoma Cell Model: The Impact of Oxygen Level on Cell Phenotype.

Treatment of osteosarcoma is hampered by tumor hypoxia and requires alternative approaches. Although the CCL2-CCR2 axis is indispensable in tumor-induced inflammation and angiogenesis, its blockade has not been effective to date. This study aimed to characterize how CCR2 inhibition affects the crosstalk of osteosarcoma cells with immune cells to better delineate tumor resistance mechanisms that help withstand such treatment. In this study, 143B cells were exposed to healthy donor PBMC supernatants in a transwell assay lacking direct cell-to-cell contact and subjected to different oxygen concentrations. In addition, mice bearing orthotopic 143B tumors were subjected to CCR2 antagonist treatment. Our findings show that hypoxic conditions alter cytokine and cancer- related protein expression on cells and impair CCR2 antagonist effects in the experimental osteosarcoma model. CCL2-CCR2 axis blockade in the 143B xenografts, which are positive for hypoxia marker CAIX, did not slow 143B tumor growth or metastasis but altered tumor microenvironment by VEGFR downregulation and shift in the CD44-positive cell population towards high CD44 expression. This study highlights differential responses of tumor cells to CCR2 antagonists in the presence of different oxygen saturations and expands our knowledge of compensatory mechanisms leading to CCL2-CCR2 treatment resistance.

Dysregulated airway epithelial antiviral immunity in Down Syndrome impairs type III IFN response and amplifies airway inflammation during RSV infection.

Trisomy 21 (TS21), also known as Down syndrome (DS), increases pediatric mortality risk from respiratory syncytial virus (RSV) by nine-fold, yet its underlying immunological basis remains unclear. Here, we investigated RSV-induced immunological responses in TS21 airway epithelial cells (AECs), the primary site of respiratory virus entry and host defense. TS21 AECs exhibit hyperactive interferon (IFN) signaling and reduced RSV infectivity, but they also show impaired type-III IFN responses during viral infection. Furthermore, TS21 AECs demonstrate heightened production of proinflammatory mediators CXCL5 and CXCL10 both before and after RSV exposure. Infants with DS suffering from severe viral bronchiolitis demonstrate dysregulated airway immune responses in vivo, characterized by diminished type-III IFN levels and increased CXCL5/CXCL10 secretion. Our results indicate that RSV severity in DS is not due to impaired viral control but to dysregulated airway proinflammatory responses, offering new therapeutic opportunities to mitigate the severity of RSV infection in children with DS.

Cancer Therapies Targeting Cellular Metabolism.

Cancer is caused by mutations that drive aberrant growth, proliferation, and invasion, thus overriding regulatory mechanisms that normally link these processes to organismal needs and cellular physiology. This imposes demands for the production of energy and biomass and for survival in microenvironments that are often nonphysiologic and nutrient-poor, which are met by rewiring of cellular metabolism. The resultant dependence of tumor cells on altered metabolism can induce sensitivity to specific metabolic perturbations that can be exploited for cancer therapy. Some cancers are caused by mutations that impart a novel function to metabolic enzymes, leading to the production of a tumor-promoting metabolite that is dispensable in normal cells, representing an ideal therapeutic target. Tumors can also exploit metabolic regulation of cellular immunity to evade antitumor immune responses, and deciphering this biology has revealed potential targets for therapeutic intervention. Here, we discuss a number of illustrative examples highlighting the therapeutic potential and the challenges of targeting metabolism for cancer therapy.

Regulation of Oxygen in the Tumor Microenvironment Synergizes with Immunotherapy to Suppress Tumor Progression

Hypoxia represents a crucial characteristic of the tumor microenvironment, which is closely related to cell proliferation, angiogenesis, and metabolic responses. These factors will further promote tumor progression, increase tumor invasion, and enhance tumor metastasis potential. A hypoxic microenvironment will also inhibit the activity of infiltrated immune cells in the tumor microenvironment, leading to the failure of cancer immunotherapy. Additionally, the hypoxic tumor microenvironment contributes to resistance to conventional therapies and leads to unfavorable prognoses. This review discusses advancements in strategies aimed at ameliorating tumor hypoxia within the microenvironment and modulating immune cell responses against tumors.

Peptide Antigen‐Surfaced TLR9‐Adjuvanting Nanovaccines as a Versatile Therapeutic Modality to Eradicate Acute Myeloid Leukemia

AbstractPeptide antigen vaccines constitute a promising clinical strategy for treating cancer patients. However, their anticancer immune response remains modest because of the suboptimal presentation of peptide antigens and/or insufficient activation of antigen‐presenting cells (APCs). The development of therapeutic vaccines for acute myeloid leukemia (AML) poses an even greater challenge because AML cells disseminate throughout the body. In this study, the peptide antigen‐surfaced TLR9‐adjuvanting nanovaccines (PASTA‐NV), which display multiple Wilm's tumor 1 (WT1) peptides on their surface and encapsulate CpG ODN adjuvants within their watery interior to simultaneously increase antigen presentation and APC activation/proliferation, are reported. Interestingly, systemic administration of PASTA‐NV induces strong cellular and humoral anticancer immune responses in orthotopic murine AML MLL‐AF9 models. When combined with an anti‐CTLA4 antibody, PASTA‐NV achieves complete regression of AML in mouse models and establishes durable anti‐AML immunity, effectively resisting rechallenge with leukemic cells. PASTA‐NV provides a new and general avenue to induce robust and specific anticancer immunity, which has the potential to revive peptide antigen‐based nanovaccines for tumor therapy.

Hypochlorous acid induces a redox-dependent growth of C2C12 myoblasts.

Hypochlorous acid (HOCl) is a reactive chlorine species generated by the enzyme myeloperoxidase present in phagocytes. HOCl plays a vital role in inflammation and has been linked to tissue regeneration through redox signalling, however, the relevant evidence is rather scarce. The present investigation aimed to study the effects of HOCl on the growth of C2C12 myoblasts and its association with alterations of cellular redox profile. C2C12 cells were incubated for 10 min, 1 h and 24 h with a wide range of HOCl concentrations (628 pM - 4 M). Cell survival was increased when cells were incubated with HOCl concentrations between 6.28 μM and 628 μM, which are encountered in biological systems. Intriguingly, after a 10 min-incubation with 3 mM of HOCl, the highest cell growth was observed through a redox-related mechanism, as indicated by the decrease of the levels of reactive oxygen species and the enhanced levels of reduced glutathione measured by flow cytometry. The in vitro model created herein simulates the in vivo inflammatory and regeneration response of muscle cells and can putatively give mechanistic answers about the contribution of HOCl to muscle regeneration.

Transcriptome analysis of infected human macrophages between strains of Brucella melitensis and an omp31 mutant.

Brucella spp. are small aerobes non-motile Gram-negative coccobacilli that act as facultative intracellular pathogens responsible for zoonotic infections. B. melitensis can survive and replicate within host macrophages, the molecular phenomena of this host: pathogen interaction remain totally unknown. The aim of this work was to evaluate the differences in the response between human macrophages infected with different B. melitensis strains. Comparison of transcriptome data was carried out for identifying differentially expressed genes among different strain infection. We evaluated the THP-1 macrophage molecular response at early stages of infection to different strains of Brucella melitensis (B. melitensis wild- type 133 (BM133), B. melitensis ATCC 23456 (BM16M) and a B. melitensis 133 omp31 mutant (LVM31)). Our analysis revealed intriguing differences in the host cell response to two virulent strains (BM16M and BM133), infection with BM16M led to an over-expression of anti-inflammatory pathways, such as cAMP signaling and PI3K-Akt pathway, and down regulation of inflammatory pathways involving IL1A and IL10 compared to BM133. Mutant strain BMLVM31 induced an activation of the apoptotic process and the absence of Omp31, impaired the inhibition of CASP1 and CASP9 expression. Additionally, the mutation of BMLV31 impairs the evasion of cathepsin D in early stages of the infection. These findings shed light on the intricate molecular interactions between B. melitensis strains and human macrophages, providing valuable insights for understanding the pathogenesis of brucellosis.

Outcome of SARS-CoV-2 reinfection depends on genetic background in female mice

Antigenically distinct SARS-CoV-2 variants increase the reinfection risk for vaccinated and previously exposed population due to antibody neutralization escape. COVID-19 severity depends on many variables, including host immune responses, which differ depending on genetic predisposition. To address this, we perform immune profiling of female mice with different genetic backgrounds –transgenic K18-hACE2 and wild-type 129S1– infected with the severe B.1.351, 30 days after exposure to the milder BA.1 or severe H1N1. Prior BA.1 infection protects against B.1.351-induced morbidity in K18-hACE2 but aggravates disease in 129S1. H1N1 protects against B.1.351-induced morbidity only in 129S1. Enhanced severity in B.1.351 re-infected 129S1 is characterized by an increase of IL-10, IL-1β, IL-18 and IFN-γ, while in K18-hACE2 the cytokine profile resembles naïve mice undergoing their first viral infection. Enhanced pathology during 129S1 reinfection cannot be attributed to weaker adaptive immune responses to BA.1. Infection with BA.1 causes long-term differential remodeling and transcriptional changes in the bronchioalveolar CD11c+ compartment. K18-hACE2 CD11c+ cells show a strong antiviral defense expression profile whereas 129S1 CD11c+ cells present a more pro-inflammatory response upon restimulation. In conclusion, BA.1 induces cross-reactive adaptive immune responses in K18-hACE2 and 129S1, but reinfection outcome correlates with differential CD11c+ cells responses in the alveolar space.

Single-cell sequencing reveals PTX3 involvement in ovarian cancer metastasis

BackgroundPentraxin 3 (PTX3) has been associated with the development and progression of various malignant tumors. However, its roles and the mechanisms underlying its involvement in ovarian cancer (OC) peritoneal metastasis remain unclear.MethodsSingle-cell RNA sequencing (scRNA-seq) and immunohistochemistry (IHC) were conducted to determine the expression profiles, potential functionalities, and underlying mechanisms of PTX3 within the context of OC. To assess the proliferative response of OC cells, we utilized both EdU (5-ethynyl-2’ -deoxyuridine) and CCK8 assays. The role of PTX3 in facilitating cell migration and invasion was quantified through the use of Transwell assays. The protein expression levels were meticulously analyzed via Western blotting. Furthermore, to explore the interactions between proteins, we conducted immunofluorescence (IF) staining and co-immunoprecipitation (Co-IP) experiments. To determine the factors responsible for the upregulation of PTX3, we performed both coculture and suspension assays, providing a comprehensive approach to understanding the regulatory mechanisms involved.ResultsThis study confirmed, for the first time, that the expression of PTX3 in OC metastatic lesions is greater than that in primary lesions and that tumor cells with high PTX3 expression have greater metastatic ability. PTX3 can activate the EMT and NF-κB signaling pathways in OC cells and can interact with the TLR4 and CD44 receptors in OC cells. Additionally, PTX3’s modulation of the EMT and NF-κB pathways is partially dependent on its interaction with TLR4. Furthermore, this study revealed the intercellular regulatory network related to PTX3 in OC cells via bioinformatic analysis. High levels of PTX3 in OC cells potentially enhance the attraction of dendritic cells (DCs) and CD4 + T cells while diminishing the recruitment of B cells and CD8 + T cells. Finally, this study indicated that PTX3 upregulation was driven by multiple factors, including specific transcription factors (TFs) and modifications within the tumor microenvironment (TME).ConclusionsOur research revealed the contribution of PTX3 to the peritoneal dissemination process in OC patients, identifying a novel potential biomarker and therapeutic target for this disease.

Transcriptomic and epigenomic profiling reveals altered responses to diesel emissions in Alzheimer's disease both in vitro and in population-based data.

Studies have correlated living close to major roads with Alzheimer's disease (AD) risk. However, the mechanisms responsible for this link remain unclear. We exposed olfactory mucosa (OM) cells of healthy individuals and AD patients to diesel emissions (DE). Cytotoxicity of exposure was assessed, mRNA, miRNA expression, and DNA methylation analyses were performed. The discovered altered pathways were validated using data from the human population-based Rotterdam Study. DE exposure resulted in an almost four-fold higher response in AD OM cells, indicating increased susceptibility to DE effects. Methylation analysis detected different DNA methylation patterns, revealing new exposure targets. Findings were validated by analyzing data from the Rotterdam Study cohort and demonstrated a key role of nuclear factor erythroid 2-related factor 2 signaling in responses to air pollutants. This study identifies air pollution exposure biomarkers and pinpoints key pathways activated by exposure. The data suggest that AD individuals may face heightened risks due to impaired cellular defenses. Healthy and AD olfactory cells respond differently to DE exposure. AD cells are highly susceptible to DE exposure. The NRF2 oxidative stress response is highly activated upon air pollution exposure. DE-exposed AD cells activate the unfolded protein response pathway. Key findings are also confirmed in a population-based study.

TNF signaling mediates lipopolysaccharide-induced lung epithelial progenitor cell responses in mouse lung organoids

Bacterial respiratory infections are a major global health concern, often leading to lung injury and triggering lung repair mechanisms. Endogenous epithelial progenitor cells are crucial in this repair, yet the mechanisms remain poorly understood. This study investigates the response of lung epithelial progenitor cells to injury induced by lipopolysaccharide (LPS), a component of gram-negative bacteria, focusing on their regulation during lung repair. Lung epithelial cells (CD31-CD45-Epcam+) from wild-type and tumor necrosis factor (TNF) receptor 1/2 knock-out mice were co-cultured with wild-type fibroblasts. Organoid numbers and size were measured after 14 days of exposure to 100 ng/mL LPS. Immunofluorescence was used to assess differentiation (after 14 days), RNA sequencing analyzed gene expression changes (after 72 hours), and MTS assay assessed proliferative effects of LPS on individual cell types (after 24 hours). LPS treatment increased the number and size of wild-type lung organoids and promoted alveolar differentiation, indicated by more SPC+ organoids. RNA sequencing revealed upregulation of inflammatory and fibrosis-related markers, including Cxcl3, Cxcl5, Ccl20, Mmp13, and Il33, and enrichment of TNF-α signaling and epithelial-mesenchymal transition pathways. TNF receptor 1 deficiency inhibited LPS-induced progenitor cell activation and organoid growth. In conclusion, LPS enhances lung epithelial progenitor cell proliferation and differentiation via TNF receptor 1 signaling, highlighting potential therapeutic targets for bacterial lung injury.

Immune responses drive chorioretinitis and retinal pathology after neonatal CMV infection.

Human cytomegalovirus (CMV) causes a common congenital infection leading to long-term neurological impairments including brain, cochlear, and ocular pathology. Infection of newborn mice with murine (M)CMV is an established model of neuropathology caused by congenital CMV infection, with recent work suggesting that brain pathology may be driven by immune responses. In the eye, however, CMV retinitis is thought to result from virus-driven necrosis in the absence of T cell responses. We found that MCMV infection of newborn mice recapitulates human eye disease after congenital CMV infection, including focal chorioretinitis, inflamed vasculature, and disrupted blood-retinal barriers. Moreover, infection drove extensive T cell infiltration of the retina and marked gliosis. Blocking immune responses generally, or via targeting the chemokine receptor CXCR3, did not exacerbate retinal disease but instead prevented pathology despite retinal MCMV infection. Thus, our data establish this model for studies of congenital retinal disease and show that the immune system drives pathology in the neonatal eye after MCMV infection.

Immune status in women after COVID-19 living in the subarctic region

Background. T cell immunity is known to play a central role in controlling SARSCoV- 2 (severe acute respiratory syndrome-related coronavirus 2) infection, so it is critical to understand its role in recovery from COVID-19 (coronavirus disease 2019), especially in the unfavorable conditions of the North.The aim. To assess the immune status of women after COVID-19 living in the subarctic region of the Russian Federation.Materials and methods. We examined a total of 50 women aged 36–46 years, including 38 vaccinated women 6 and 12 months after recovery from COVID-19 and 12 women (control group) who had not had COVID-19 and had not been vaccinated (Arkhangelsk region). We studied leukocyte count, leukogram and lymphocyte phenotypes (CD5+, CD8+, CD10+, CD16+, CD95+).Results. The contribution of monocytes to the formation of the adaptive immune response among female residents of the subarctic region decreases in those who have had mild COVID-19 and increases in those who have had moderate COVID-19. It was found that the imbalance in the formation of the immune response after COVID-19 is formed due to innate immunity (CD16+, neutrophils (r = 0.89; p < 0.001)), affects the adaptive immunity and depends on the severity of the disease and the time since recovery. The most significant contribution to the formation of the adaptive immune response was established due to cellmediated cytotoxicity (CD8+, CD16+) and the activity of apoptotic processes (CD95+) both after 6 and 12 months and does not depend on the severity of the COVID-19. The adaptive immune response formed 6 months after COVID-19 with a high level of cell-mediated cytotoxicity and activity of apoptotic processes prevailing over lymphoproliferation persists after 12 months, which indicates a tense state of immune homeostasis.Conclusion. In the examined women who had recovered from COVID-19, cellmediated cytotoxicity (CD8+, CD16+) is associated with the activation of the monocyte system, has a prolonged effect of up to 12 months and depends on the severity of the previous COVID-19 in 66.7–90 % of cases.