Preclinical evidence in the assembly of mammalian SWI/SNF complexes: Epigenetic insights and clinical perspectives in human lung disease therapy.

The SWI/SNF complex, also known as the BRG1/BRM-associated factor (BAF) complex, represents a critical regulator of chromatin remodeling mechanisms in mammals. It is alternatively referred to as mSWI/SNF and has been suggested to be imbalanced in human disease compared with human health. Three types of BAF assemblies associated with it have been described, including (1) canonical BAF (cBAF), (2) polybromo-associated BAF (PBAF), and (3) non-canonical BAF (ncBAF) complexes. Each of these BAF assemblies plays a role, either functional or dysfunctional, in governing gene expression patterns, cellular processes, epigenetic mechanisms, and biological processes. Recent evidence increasingly links the dysregulation of mSWI/SNF complexes to various human non-malignant lung chronic disorders and lung malignant diseases. This review aims to provide a comprehensive general state-of-the-art and a profound examination of the current understanding of mSWI/SNF assembly processes, as well as the structural and functional organization of mSWI/SNF complexes and their subunits. In addition, it explores their intricate functional connections with potentially dysregulated transcription factors, placing particular emphasis on molecular and cellular pathogenic processes in lung diseases. These processes are reflected in human epigenome aberrations that impact clinical and therapeutic levels, suggesting novel perspectives on the diagnosis and molecular therapies for human respiratory diseases.

STAT6 controls the stability and suppressive function of regulatory T cells.

Signal transducer and activator of transcription 6 (STAT6) promotes tumorigenesis by decreasing the Forkhead box P3+ (Foxp3+) cell frequency allowing for the infiltration of inflammatory cells during the early stages of colitis-associated cancer (CAC). In this study, we dissected the role of STAT6 in the generation of inducible in vitro regulatory T cells (iTregs) and peripheral in vivo Tregs (pTregs) under inflammatory conditions. In in vitro assays, when STAT6 was lacking, iTregs preserved a stable phenotype and expressed high levels of Foxp3 and CD25 during long expansion periods, even in the presence of IL-6. This effect was associated with increased in vitro suppressive ability, over-expression of programmed death-1 (PD-1), CTLA-4, and Foxp3, and decreased IFN-γ expression. Furthermore, iTregs developed during STAT6 deficiency showed a higher demethylation status for the FOXP3 Treg-specific demethylated region (TSDR), coupled with lower DNA methyltransferase 1 (DNMT1) mRNA expression, suggesting that STAT6 may lead to Foxp3 silencing. Using a mouse model of CAC, the STAT6-/- pTregs expressed a more activated phenotype at the intestine, had higher suppressive capacity, and expressed more significant levels of PD-1 and latency-associated peptide of TGF-ß (LAP) associated with their ability to attenuate tumor development. These data suggest that STAT6 signaling impairs the induction, stability, and suppressive capacity of Tregs developed in vitro or in vivo during gut inflammation.

Improved biodistribution and enhanced immune response of subunit vaccine using a nanostructure formed by self-assembly of ascorbyl palmitate.

New adjuvant strategies are needed to improve protein-based subunit vaccine immunogenicity. We examined the potential to use nanostructure of 6-O-ascorbyl palmitate to formulate ovalbumin (OVA) protein and an oligodeoxynucleotide (CpG-ODN) (OCC). In mice immunized with a single dose, OCC elicited an OVA-specific immune response superior to OVA/CpG-ODN solution (OC). Rheological studies demonstrated OCC's self-assembling viscoelastic properties. Biodistribution studies indicated that OCC prolonged OVA and CpG-ODN retention at injection site and lymph nodes, reducing systemic spread. Flow-cytometry assays demonstrated that OCC promoted OVA and CpG-ODN co-uptake by Ly6ChiCD11bhiCD11c+ monocytes. OCC and OC induced early IFN-γ in lymph nodes, but OCC led to higher concentration. Conversely, mice immunized with OC showed higher serum IFN-γ concentration compared to those immunized with OCC. In mice immunized with OCC, NK1.1+ cells were the IFN-γ major producers, and IFN-γ was essential for OVA-specific IgG2c switching. These findings illustrate how this nanostructure improves vaccine's response.

Influence of low insertion torque values on survival rate of immediately loaded dental implants: A systematic review and meta-analysis.

AIM: The aim was to systematically evaluate the effect of low insertion torque values on the survival rate of immediately loaded dental implants. MATERIALS AND METHODS: The protocol was registered with PROSPERO (ID CRD42020189499). An electronic search was performed in PubMed, Embase, Web of Science, and Cochrane Central Register of Controlled Trials until June 2022 in English and Spanish. Studies analysing the failure or survival rate of immediately loaded dental implants according to different insertion torque values were included. RESULTS: Five-hundred seventy-three articles were assessed for eligibility, of which seven articles, four randomized clinical trials (RCTs), one controlled clinical trial, and two prospective case series studies were included in the qualitative analysis. The RCTs were classified as having low risk of bias and the non-RCTs as having moderate and serious risk of bias. The mean survival rate for implants with low insertion toque (≤35 Ncm) was 96% (p > .001, 95% confidence interval [CI]: 0.91-0.98) and that for implants with medium or high insertion torque (>35 Ncm) was 92% (p > .001, 95% CI: 0.86-0.96) (incidence rate ratio [IRR] = 1.05, 95% CI: 0.79-1.39, p = .175, I2  = 0.0%). Splinted implants with insertion torque >20 Ncm and single implants with insertion torque >35 Ncm had a higher survival rate than implants with lower insertion torque values (IRR = 1.05, 95% CI: 0.78-1.43, p = .956, I2  = 0.0%, and RR = 0.92, 95% CI: 0.48-1.75, p = .799, I2  = 0.0%, respectively). Different insertion torque values achieved equivalent outcomes. The mean follow-up was 24 months. CONCLUSIONS: Low insertion torque values have no significant effect on survival rates of immediate loading implants at a mean follow-up of 24 months.

Mexican Colorectal Cancer Research Consortium (MEX-CCRC): Etiology, Diagnosis/Prognosis, and Innovative Therapies.

In 2013, recognizing that Colorectal Cancer (CRC) is the second leading cause of death by cancer worldwide and that it was a neglected disease increasing rapidly in Mexico, the community of researchers at the Biomedicine Research Unit of the Facultad de Estudios Superiores Iztacala from the Universidad Nacional Autónoma de México (UNAM) established an intramural consortium that involves a multidisciplinary group of researchers, technicians, and postgraduate students to contribute to the understanding of this pathology in Mexico. This article is about the work developed by the Mexican Colorectal Cancer Research Consortium (MEX-CCRC): how the Consortium was created, its members, and its short- and long-term goals. Moreover, it is a narrative of the accomplishments of this project. Finally, we reflect on possible strategies against CRC in Mexico and contrast all the data presented with another international strategy to prevent and treat CRC. We believe that the Consortium's characteristics must be maintained to initiate a national strategy, and the reported data could be useful to establish future collaborations with other countries in Latin America and the world.

Natural history of circulating miRNAs in Duchenne disease: Association with muscle injury and metabolic parameters.

OBJECTIVES: This study aimed to evaluate whether the expression of circulating dystromiRs and a group of oxidative stress-related (OS-R) miRNAs is associated with muscle injury and circulating metabolic parameters in Duchenne muscular dystrophy (DMD) patients. METHODS: Twenty-four DMD patients were included in this cross-sectional study. Clinical scales to evaluate muscle injury (Vignos, GMFCS, Brooke, and Medical Research Council), enzymatic muscle injury parameters (CPK, ALT, and AST), anthropometry, metabolic indicators, physical activity, serum dystromiRs (miR-1-3p, miR-133a-3p, and miR-206), and OS-R miRNAs (miR-21-5p, miR-31-5p, miR-128-3p, and miR-144-3p) levels were measured in ambulatory and non-ambulatory DMD patients. RESULTS: DystromiRs (except miR-1-3p) and miRNAs OS-R levels were lower (p-value <.05) in the non-ambulatory group than the ambulatory group. The expression of those miRNAs correlated with Vignos scale score (For instance, rho = -0.567, p-value <0.05 for miR-21-5p) and with other scales scores of muscle function and strength. CPK, AST, and ALT concentration correlated with expression of all miRNAs (For instance, rho = 0.741, p-value <.05 between miR-206 level and AST concentration). MiR-21-5p level correlated with glucose concentration (rho = -0.369, p-value = .038), and the miR-1-3p level correlated with insulin concentration (rho = 0.343, p-value = .05). CONCLUSIONS: Non-ambulatory DMD patients have lower circulating dystromiRs and OS-R miRNAs levels than ambulatory DMD patients. The progressive muscle injury is associated with a decrease in the expression of those miRNAs, evidencing DMD progress. These findings add new information about the natural history of DMD.

Recurrence and progression of periodontitis and methods of management in long-term care: A systematic review and meta-analysis.

AIM: To systematically review the literature to evaluate the recurrence of disease of people in long-term supportive periodontal care (SPC), previously treated for periodontitis, and determine the effect of different methods of managing recurrence. The review focused on stage IV periodontitis. MATERIALS AND METHODS: An electronic search was conducted (until May 2020) for prospective clinical trials. Tooth loss was the primary outcome. RESULTS: Twenty-four publications were retrieved to address recurrence of disease in long-term SPC. Eight studies were included in the meta-analyses for tooth loss, and three studies for disease progression/recurrence (clinical attachment level [CAL] loss ≥2 mm). For patients in SPC of 5-20 years, prevalence of losing more than one tooth was 9.6% (95% confidence interval [CI] 5%-14%), while experiencing more than one site of CAL loss ≥2 mm was 24.8% (95% CI 11%-38%). Six studies informed on the effect of different methods of managing recurrence, with no clear evidence of superiority between methods. No data was found specifically for stage IV periodontitis. CONCLUSIONS: A small proportion of patients with stage III/IV periodontitis will experience tooth loss in long-term SPC (tendency for greater prevalence with time). Regular SPC appears to be important for reduction of tooth loss. No superior method to manage disease recurrence was found.

Expression of Surfactant Protein D Distinguishes Severe Pandemic Influenza A(H1N1) from Coronavirus Disease 2019.

The differentiation between influenza and coronavirus disease 2019 (COVID-19) could constitute a diagnostic challenge during the ongoing winter owing to their clinical similitude. Thus, novel biomarkers are required to enable making this distinction. Here, we evaluated whether the surfactant protein D (SP-D), a collectin produced at the alveolar epithelium with known immune properties, was useful to differentiate pandemic influenza A(H1N1) from COVID-19 in critically ill patients. Our results revealed high serum SP-D levels in patients with severe pandemic influenza but not those with COVID-19. This finding was validated in a separate cohort of mechanically ventilated patients with COVID-19 who also showed low plasma SP-D levels. However, plasma SP-D levels did not distinguish seasonal influenza from COVID-19 in mild-to-moderate disease. Finally, we found that high serum SP-D levels were associated with death and renal failure among severe pandemic influenza cases. Thus, our studies have identified SP-D as a unique biomarker expressed during severe pandemic influenza but not COVID-19.

Periodontitis and systemic markers of neurodegeneration: A case-control study.

AIM: To investigate whether periodontitis is associated with amyloid beta (Aß) peptides and whether systemic inflammation could act as a potential mediator of this link. MATERIALS AND METHODS: A case-control study was designed including 75 patients with periodontitis (cases) and 75 age-balanced and gender-matched participants without periodontitis (controls). Full-mouth periodontal evaluation was performed in all participants. Demographic, clinical and behaviour data were also recorded. Fasting blood samples were collected, and serum levels of interleukin 6 (IL-6), high-sensitivity C-reactive protein (hs-CRP), Aß1-40 and Aß1-42 were determined. RESULTS: Cases showed higher levels of IL-6 (8.7 ± 3.2 vs. 4.8 ± 0.5 pg/ml), hs-CRP (3.3 ± 1.2 vs. 0.9 ± 0.7 mg/L), Aß1-40 (37.3 ± 6.0 vs. 30.3 ± 1.8 pg/ml) and Aß1-42 (54.5 ± 10.6 vs. 36.5 ± 10.0 pg/ml) when compared to controls (all p < .001). Diagnosis of periodontitis was statistically significantly associated with circulating Aß1-40 ( ßcoefficientadjusted  = 6.9, 95% CI: 5.4-8.3; p < .001) and Aß1-42 ( ßcoefficientadjusted  = 17.8, 95% CI: 14.4-21.3; p < .001). Mediation analysis confirmed hs-CRP and IL-6 as mediators of this association. CONCLUSIONS: Periodontitis is associated with increased peripheral levels of Aß. This finding could be explained by enhanced systemic inflammation that can be seen in patients with periodontitis.

Corrigendum to "Macrophage Migration Inhibitory Factor Promotes the Interaction between the Tumor, Macrophages, and T Cells to Regulate the Progression of Chemically Induced Colitis-Associated Colorectal Cancer".

[This corrects the article DOI: 10.1155/2019/2056085.].

Macrophage Migration Inhibitory Factor Promotes the Interaction between the Tumor, Macrophages, and T Cells to Regulate the Progression of Chemically Induced Colitis-Associated Colorectal Cancer.

Colitis-associated colorectal cancer (CRC) development has been shown to be related to chronically enhanced inflammation. Macrophage migration inhibitory factor (MIF) is an inflammatory mediator that favors inflammatory cytokine production and has chemotactic properties for the recruitment of macrophages (Møs) and T cells. Here, we investigated the role of MIF in the inflammatory response and recruitment of immune cells in a murine model of chemical carcinogenesis to establish the impact of MIF on CRC genesis and malignancy. We used BALB/c MIF-knockout (MIF-/-) and wild-type (WT) mice to develop CRC by administering intraperitoneal (i.p.) azoxymethane and dextran sodium sulfate in drinking water. Greater tumor burdens were observed in MIF-/- mice than in WT mice. Tumors from MIF-/- mice were histologically identified to be more aggressive than tumors from WT mice. The localization of MIF suggests that it is also involved in cell differentiation. The relative gene expression of il-17, measured by real-time PCR, was higher in MIF-/- CRC mice, compared to the WT CRC and healthy MIF-/- mice. Importantly, compared to the WT intestinal epithelium, lower percentages of tumor-associated Møs were found in the MIF-/- intestinal epithelium. These results suggest that MIF plays a role in controlling the initial development of CRC by attracting Møs to the tumor, which is a condition that favors the initial antitumor responses.

Epigenetics in non-small cell lung carcinomas.

OBJECTIVE: To perform a systematic review of the main epigenetic aberrations involved in non-small cell lung carcinomas' (NSCLC) diagnosis, progression, and therapeutics. MATERIALS AND METHODS: We performed a systematic review of the scientific literature on lung cancer epigenetics, focusing on NSCLC. RESULTS: Several advances in the molecular study of classical epigenetic mechanisms and massive studies of lung cancer epigenome have contributed relevant new evidence revealing that various molecular complexes are functionally influencing genetic-epigenetic and transcriptional mechanisms that promote lung tumorigenesis (initiation, promotion, and progression), and are also involved in NSCLC therapyresistance mechanisms. CONCLUSIONS: Several epigenetic complexes and mechanisms must be analyzed and considered for the design of new and efficient therapies, which could be fundamental to develop an integrated knowledge to achieve a comprehensive lung cancer personalized medicine.

OBJETIVO: Realizar una revisión sistemática y estructurada de las principales aberraciones epigenéticas involucradas en el diagnóstico, progresión y terapia del cáncer pulmonar de células no pequeñas (CPCNP). MATERIAL Y MÉTODOS: Revisión sistemática de literatura científica sobre epigenética del cáncer pulmonar del grupo CPCNP. RESULTADOS: El estudio de los diversos mecanismos epigenéticos y su impronta epigenética en el epigenoma del cáncer pulmonar han arrojado nuevas evidencias a nivel biológico, biomédico y médico-clínico del impacto que los mecanismos epigenéticotranscripcionales promueven de manera activa y reversible sobre los procesos de tumorigénesis, progresión histopatológica y mecanismos de resistencia a la terapia oncológica pulmonar. CONCLUSIONES: Deben analizarse diferentes complejos y mecanismos epigenéticos para el estudio y diseño de esquemas nuevos y eficaces de terapia epigenética, los cuales podrían ser fundamentales para desarrollar un conocimiento integral en el desarrollo de la medicina personalizada en el cáncer pulmonar del grupo CPCNP.

Mean annual attachment, bone level, and tooth loss: A systematic review.

BACKGROUND: Rate of progression of periodontitis has been used to inform the design of classifications of periodontal diseases. However, the evidence underpinning this topic is unclear and no systematic review has yet been conducted. OBJECTIVES: The focused question for this systematic review was: in adults, what is the progression of periodontitis in terms of clinical attachment loss, radiographic bone loss, and tooth loss? DATA SOURCES: Highly sensitive electronic search was conducted for published data in MEDLINE, EMBASE, LILACS, and unpublished grey literature in OpenGrey up to February 2016. Reference lists of retrieved studies for full-text screening and reviews were hand-searched for potentially eligible studies. STUDY ELIGIBILITY CRITERIA AND PARTICIPANTS: Prospective, longitudinal observational studies with follow-up of at least 12 months and presenting data on the primary outcome, change in clinical attachment level, in adults (age ≥18 years). Secondary outcomes, tooth loss and bone level change, were only assessed in studies reporting the primary outcome. Studies investigating specific disease populations or only on treated periodontitis patients were excluded. STUDY APPRAISAL AND SYNTHESIS METHODS: Risk of bias and methodology were assessed using the Newcastle-Ottawa Scale with two additional questions on security of outcome assessment. Studies were pooled by abstracting or estimating mean annual attachment or bone level change and annual tooth loss. Random effects meta-analysis was conducted with investigation of effect of potential modifiers where possible. RESULTS: A total 11,482 records were screened for eligibility; 33 publications of 16 original studies reporting on more than 8,600 participants were finally included as eligible for the review. The studies represented populations from both developing and developed economies. Mean annual attachment loss was 0.1 mm per year (95% CI 0.068, 0.132; I2  = 99%) and mean annual tooth loss was 0.2 teeth per year (95% CI 0.10, 0.33; I2  = 94%). Observational analysis of highest and lowest mean attachment change quintiles suggested substantial differences between groups with minimal annual change in the lowest quintile and an average deterioration of 0.45 mm mean attachment loss per year in the highest group. This value increased to 0.6 mm per year with periodontitis alone. There was surprisingly little effect of age or gender on attachment level change. Geographic location, however, was associated with more than three times higher mean annual attachment loss in Sri Lanka and China (0.20 mm, 95% CI 0.15, 0.27; I2  = 83%) vs North America and Europe (0.056 mm, 95% CI 0.025, 0.087; I2  = 99%) P < 0.001. LIMITATIONS: There were a limited number of studies (N = 16), high variability of design in key study components (sampling frames, included ages, data analyses), and high statistical heterogeneity that could not be explained. CONCLUSIONS: Within the limitations of the research, the data show that mean annual attachment level change varies considerably both within and between populations. Overall, the evidence does not support or refute the differentiation between forms of periodontal diseases based upon progression of attachment level change.

Secreted and Tissue miRNAs as Diagnosis Biomarkers of Malignant Pleural Mesothelioma.

Malignant pleural mesothelioma (MPM) is a rare but aggressive tumor that originates in the pleura, is diagnosed in advanced stages and has a poor prognosis. Accurate diagnosis of MPM is often difficult and complex, and the gold standard diagnosis test is based on qualitative analysis of markers in pleural tissue by immunohistochemical staining. Therefore, it is necessary to develop quantitative and non-subjective alternative diagnostic tools. MicroRNAs are non-coding RNAs that regulate essential cellular mechanisms at the post-transcriptional level. Recent evidence indicates that miRNA expression in tissue and body fluids is aberrant in various tumors, revealing miRNAs as promising diagnostic biomarkers. This review summarizes evidence regarding secreted and tissue miRNAs as biomarkers of MPM and the biological characteristics associated with their potential diagnostic value. In addition to studies regarding miRNAs with potential diagnostic value for MPM, studies that aimed to identify the miRNAs involved in molecular mechanisms associated with MPM development are described with an emphasis on relevant aspects of the experimental designs that may influence the accuracy, consistency and real diagnostic value of currently reported data.

Circulating levels of miR-150 are associated with poorer outcomes of A/H1N1 infection.

BACKGROUND: Overproduction of pro-inflammatory cytokines and chemokines is frequently associated with severe clinical manifestations in patients infected with influenza A/H1N1 virus. Micro-RNAs (miRNAs) are highly conserved small non-coding RNA molecules that post-transcriptionally regulate gene expression and are potential biomarkers and therapeutic targets in different inflammatory conditions. METHODS: We studied the circulating and miRNA profiles in critically ill A/H1N1 patients, A/H1N1 patients with milder disease, asymptomatic housemates and healthy controls. Cytokine, chemokine and growth factors that were potential targets of differentially expressed miRNAs were assessed. Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway enrichment and interactome analysis of these miRNAs were also performed. RESULTS: Critically ill patients exhibited a significant over-expression of circulating miR-150 (p<0.005) when compared to patients with milder disease. miR-29c, miR-145 and miR-22 were differentially expressed in patients with severe A/H1N1 disease whereas miR-210, miR-126 and miR-222 were downregulated in individuals exposed to the A/H1N1 virus. Significant correlations (p<0.05) between circulating levels of miR-150 with IL-1ra, IL-2, IL-6, CXCL8, IFN-γ, CXCL10 and G-CSF were detected, particularly in critically ill patients. CONCLUSION: The up-regulation of miR-150 is associated with poorer outcomes of A/H1N1 infection. The differential expression of miRNAs related with immune processes in severe A/H1N1 disease supports the potential role of these miRNAs as biomarkers of disease progression.

Serum surfactant protein D (SP-D) is a prognostic marker of poor outcome in patients with A/H1N1 virus infection.

INTRODUCTION: Surfactant protein D (SP-D) plays an important role in the innate responses against pathogens and its production is altered in lung disorders. METHODS: We studied the circulating levels of SP-D in 37 patients with acute respiratory distress syndrome due to the A/H1N1 virus infection and in 40 healthy controls. Cox logistic regression models were constructed to explore the association of SP-D levels and risk of death. RESULTS: Mortality rate after a 28-day was 32.42 %. Significant higher levels of SP-D were detected in A/H1N1 patients with fatal outcome (p < 0.05). After adjusting for confounding variables, levels of SP-D ≥250 ng/mL were associated with increased the risk of death (HR = 8.27, 95 % CI 1.1-64.1, p = 0.043). CONCLUSIONS: Our results revealed that higher circulating levels of SP-D are associated with higher mortality risk in critically ill A/H1N1 patients. SP-D might be a predictive factor of poor outcomes in viral pneumonia.

Obesity and pro-inflammatory mediators are associated with acute kidney injury in patients with A/H1N1 influenza and acute respiratory distress syndrome.

BACKGROUND: The obesity has been shown to increase the severity of A/H1N1 infection and the development of acute respiratory distress syndrome (ARDS) and organ involvement. METHODS: Circulating levels of C-peptide, insulin, glucagon, leptin, acute phase reactants (procalcitonin, C-reactive protein, tissue plasminogen activator, and serum amyloids A and P), were measured in samples from 32 critically ill patients with A/H1N1 virus infection, 17 of whom had ARDS complicated by acute kidney injury (AKI) and 15 of whom had ARDS but did not develop AKI. RESULTS: Patients with ARDS and AKI (ARDS/AKI) had higher BMI and higher levels of C-peptide, insulin, leptin, procalcitonin and serum amyloid A compared to those ARDS patient who did not develop AKI. Adjusting for confounding variables using logistic regression analysis, higher levels of C-peptide (>0.75 ng/mL) (OR=64.8, 95% CI = 2.1-1980, p = 0.0006) and BMI>30 Kg/m(2) (OR = 42.0, 95% CI = 1.2-1478, p = 0.04) were significantly associated with the development of AKI in ARDS patients. CONCLUSION: High levels of C-peptide and BMI>30 kg/m(2) were associated with the development of AKI in ARDS patients due to A/H1N1 infection. These metabolic/obesity indicators, together with the profiles of pro-inflammatory acute phase proteins, may be important links between obesity and poor outcomes in A/H1N1 09 infection.

Targeted Demethylation of FOXP3-TSDR Enhances the Suppressive Capacity of STAT6-deficient Inducible T Regulatory Cells.

In vitro induced T regulatory cells (iTregs) are promising for addressing inflammation-driven diseases. However, current protocols for the generation and expansion of iTregs fail to induce extensive demethylation of the Treg-specific demethylated region (TSDR) within the FOXP3 gene, recognized as the master regulator for regulatory T cells (Tregs). This deficiency results in the rapid loss of Foxp3 expression and an unstable regulatory phenotype. Nevertheless, inhibition of STAT6 signaling effectively stabilizes Foxp3 expression in iTregs. Thus, this study aimed to develop a protocol combining epigenetic editing with STAT6 deficiency to improve iTregs' ability to maintain stable suppressive function and a functional phenotype. Our findings demonstrate that the combination of STAT6 deficiency (STAT6-/-) with targeted demethylation of the TSDR using a CRISPR-TET1 tool leads to extensive demethylation of FOXP3-TSDR. Demethylation in STAT6-/- iTregs was associated with enhanced expression of Foxp3 and suppressive markers such as CTLA-4, PD-1, IL-10, and TGF-ß. Furthermore, the edited STAT6-/- iTregs exhibited an increased capacity to suppress CD8+ and CD4+ lymphocytes and could more efficiently impair Th1-signature gene expression compared to conventional iTregs. In conclusion, the deactivation of STAT6 and TSDR-targeted demethylation via CRISPR-TET1 is sufficient to induce iTregs with heightened stability and increased suppressive capacity, offering potential applications against inflammatory and autoimmune diseases.