Evolution and molecular interactions of major histocompatibility complex (MHC)-G, -E and -F genes.

Classical HLA (Human Leukocyte Antigen) is the Major Histocompatibility Complex (MHC) in man. HLA genes and disease association has been studied at least since 1967 and no firm pathogenic mechanisms have been established yet. HLA-G immune modulation gene (and also -E and -F) are starting the same arduous way: statistics and allele association are the trending subjects with the same few results obtained by HLA classical genes, i.e., no pathogenesis may be discovered after many years of a great amount of researchers' effort. Thus, we believe that it is necessary to follow different research methodologies: (1) to approach this problem, based on how evolution has worked maintaining together a cluster of immune-related genes (the MHC) in a relatively short chromosome area since amniotes to human at least, i.e., immune regulatory genes (MHC-G, -E and -F), adaptive immune classical class I and II genes, non-adaptive immune genes like (C2, C4 and Bf) (2); in addition to using new in vitro models which explain pathogenetics of HLA and disease associations. In fact, this evolution may be quite reliably studied during about 40 million years by analyzing the evolution of MHC-G, -E, -F, and their receptors (KIR-killer-cell immunoglobulin-like receptor, NKG2-natural killer group 2-, or TCR-T-cell receptor-among others) in the primate evolutionary lineage, where orthology of these molecules is apparently established, although cladistic studies show that MHC-G and MHC-B genes are the ancestral class I genes, and that New World apes MHC-G is paralogous and not orthologous to all other apes and man MHC-G genes. In the present review, we outline past and possible future research topics: co-evolution of adaptive MHC classical (class I and II), non-adaptive (i.e., complement) and modulation (i.e., non-classical class I) immune genes may imply that the study of full or part of MHC haplotypes involving several loci/alleles instead of single alleles is important for uncovering HLA and disease pathogenesis. It would mainly apply to starting research on HLA-G extended haplotypes and disease association and not only using single HLA-G genetic markers.

TGFB1 polymorphisms and TGF-ß1 plasma levels identify gastric adenocarcinoma patients with lower survival rate and disseminated disease.

TGF-ß1 is involved in tumour growth. Four TGFB1 SNPs and TGF-ß1 production by stimulated PBMC were determined in seventy-eight gastric adenocarcinoma patients. In addition, TGF-ß1 levels were measured in the plasma of further thirty patients. rs1800471-G/C genotype was prevalent in patients (20.7%) compared to controls (8.4%), as it also was the rs1800468 SNP-G/A genotype in stage IV patients (20.7%) compared to stage I, II and III patients, combined (10.3%). Conversely, the T/T rs1800469 SNP-T/T genotype was absent in the former group and present in 19.0% in the latter. Furthermore, the rs1800469-C/rs1800470-T (CT) haplotype was found in 15.0% of stage IV patients as compared to 3.0% of the remaining patients (3.0%) and also identifies patients with worse five-year life expectancy (P = .03). TGF-ß1 synthesis by stimulated PBMCs was significantly lower in patients with the risk SNPs or haplotype, compared to the alternative genotype. Finally, TGF-ß1 plasma levels were lower in patients with worse life expectancy. Analysis of TGFB1 SNPs and measurement of plasma TGF-ß1 levels serves to identify patients at risk of developing a more aggressive disease.

HLA-G in Mayas from Yucatan: An evolutionary approach.

HLA-G allele frequencies were studied in Yucatán (Mexico) Maya Amerindians by a direct exon DNA sequencing technique. It is described that Mayas are probably one of the first populations together with Olmecs that populated Meso America and that important HLA genetic differences between Mexican and Guatemalan Mayas support that Maya languages were imposed to several neighbouring Amerindian groups. HLA-G*01:01:02, HLA-G*01:01:01 and HLA-G*01:04:01 are the most frequent alleles in this population. It is remarkable that HLA-G*01:05N allele was not found in the population in accordance with similar results found in another Amerindians. Also, protein allele HLA-G*01:04 frequency is found not to differ to those found in another far or close living Amerindians in contrast to other World populations. It seems that while high HLA-G*01:05N frequency is found in Iran and Middle East populations, probably where this allele appeared within an ancestral HLA-A*19 group of alleles haplotype and it is maintained by unknown evolutionary forces, Amerindians do not have a high frequency because a founder effect or because required natural evolutionary forces do not exist in America. Finally, we believe useful to study HLA-G evolution for its physiopathology understanding in addition to the many papers on statistics on HLA-G and in vitro models that are yearly published.

New insights into the genetic component of non-infectious uveitis through an Immunochip strategy.

BACKGROUND: Large-scale genetic studies have reported several loci associated with specific disorders involving uveitis. Our aim was to identify genetic risk factors that might predispose to uveitis per se, independent of the clinical diagnosis, by performing a dense genotyping of immune-related loci. METHODS: 613 cases and 3693 unaffected controls from three European case/control sets were genotyped using the Immunochip array. Only patients with non-infectious non-anterior uveitis and without systemic features were selected. To perform a more comprehensive analysis of the human leucocyte antigen (HLA) region, SNPs, classical alleles and polymorphic amino acid variants were obtained via imputation. A meta-analysis combining the three case/control sets was conducted by the inverse variance method. RESULTS: The highest peak belonged to the HLA region. A more detailed analysis of this signal evidenced a strong association between the classical allele HLA-A*2902 and birdshot chorioretinopathy (p=3.21E-35, OR=50.95). An omnibus test yielded HLA-A 62 and 63 as relevant amino acid positions for this disease. In patients with intermediate and posterior uveitis, the strongest associations belonged to the rs7197 polymorphism, within HLA-DRA (p=2.07E-11, OR=1.99), and the HLA-DR15 haplotype (DRB1*1501: p=1.16E-10, OR=2.08; DQA1*0102: p=4.37E-09, OR=1.77; DQB1*0602: p=7.26E-10, OR=2.02). Outside the HLA region, the MAP4K4/IL1R2 locus reached statistical significance (rs7608679: p=8.38E-07, OR=1.42). Suggestive associations were found at five other loci. CONCLUSIONS: We have further interrogated the association between the HLA region and non-infectious non-anterior uveitis. In addition, we have identified a new non-HLA susceptibility factor and proposed additional risk loci with putative roles in this complex condition.

Pro-inflammatory self-reactive T cells are found within murine TCR-αß(+) CD4(-) CD8(-) PD-1(+) cells.

TCR-αß(+) double negative (DN) T cells (CD3(+) TCR-αß(+) CD4(-) CD8(-) NK1.1(-) CD49b(-) ) represent a minor heterogeneous population in healthy humans and mice. These cells have been ascribed pro-inflammatory and regulatory capacities and are known to expand during the course of several autoimmune diseases. Importantly, previous studies have shown that self-reactive CD8(+) T cells become DN after activation by self-antigens, suggesting that self-reactive T cells may exist within the DN T-cell population. Here, we demonstrate that programmed cell death 1 (PD-1) expression in unmanipulated mice identifies a subset of DN T cells with expression of activation-associated markers and a phenotype that strongly suggests they are derived from self-reactive CD8(+) cells. We also found that, within DN T cells, the PD-1(+) subset generates the majority of pro-inflammatory cytokines. Finally, using a TCR-activation reporter mouse (Nur77-GFP), we confirmed that in the steady-state PD-1(+) DN T cells engage endogenous antigens in healthy mice. In conclusion, we provide evidence that indicates that the PD-1(+) fraction of DN T cells represents self-reactive cells.

Programmed cell death 1 and Helios distinguish TCR-αß+ double-negative (CD4-CD8-) T cells that derive from self-reactive CD8 T cells.

TCR-αß(+) double-negative (DN; CD4(-)CD8(-)) T cells represent a poorly understood cellular subset suggested to contribute to the pathogenesis of the autoimmune disease systemic lupus erythematosus. DN T cells have been proposed to derive from CD8(+) cells. However, the conditions that govern the loss of CD8 expression after Ag encounter are unknown. In this study, we tracked the fate of CD8 T cells from transgenic TCR mice exposed to their cognate Ags as self or in the context of infection. We demonstrate that CD8 T cells lose CD8 expression and become DN only when cognate Ag is sensed as self. This process is restricted to tissues where the Ag is present. We also show that DN T cells derived from self-reactive CD8 cells express the inhibitory molecules PD-1 and Helios. These molecules identify a subset of DN T cells in normal mice. A similar population expands when CD8 T cells from repertoires enriched in self-reactive cells (Aire-deficient) are transferred into cognate hosts. Collectively, our data suggest that a subset of DN T cells, identified by the expression of PD-1 and Helios, represent self-reactive cells. Our results provide an explanation for the origin of DN T cells and introduce CD8 loss as a process associated with self-Ag encounter.

Crk adaptors negatively regulate actin polymerization in pedestals formed by enteropathogenic Escherichia coli (EPEC) by binding to Tir effector.

Infections by enteropathogenic Escherichia coli (EPEC) cause diarrhea linked to high infant mortality in developing countries. EPEC adheres to epithelial cells and induces the formation of actin pedestals. Actin polymerization is driven fundamentally through signaling mediated by Tir bacterial effector protein, which inserts in the plasma membrane of the infected cell. Tir binds Nck adaptor proteins, which in turn recruit and activate N-WASP, a ubiquitous member of the Wiskott-Aldrich syndrome family of proteins. N-WASP activates the Arp2/3 complex to promote actin polymerization. Other proteins aside from components of the Tir-Nck-N-WASP pathway are recruited to the pedestals but their functions are unknown. Here we investigate the function of two alternatively spliced isoforms of Crk adaptors (CrkI/II) and the paralog protein CrkL during pedestal formation by EPEC. We found that the Crk isoforms act as redundant inhibitors of pedestal formation. The SH2 domain of CrkII and CrkL binds to phosphorylated tyrosine 474 of Tir and competes with Nck to bind Tir, preventing its recruitment to pedestals and thereby inhibiting actin polymerization. EPEC infection induces phosphorylation of the major regulatory tyrosine in CrkII and CrkL, possibly preventing the SH2 domain of these proteins from interacting with Tir. Phosphorylated CrkII and CrkL proteins localize specifically to the plasma membrane in contact with EPEC. Our study uncovers a novel role for Crk adaptors at pedestals, opening a new perspective in how these oncoproteins regulate actin polymerization.

Gorgan (Turkmen in Iran) HLA genetics: transplantation, pharmacogenomics and anthropology.

HLA class I and II alleles have been studied in a population from Gorgan (North East Iranian city bordering Turkmenistan). This population is composed of mainly Turkmen who speak Oghuz Turkish language. Comparison of Gorgan people HLA profile has been carried out with about 7984 HLA chromosomes from other worldwide populations; extended haplotypes and three dimension genetic distances have been calculated by using neighbor-joining and correspondence relatedness analyses. Most frequent extended HLA haplotypes show a Siberian/Mediterranean admixture and closest populations are Chuvashians (North Caspian Sea, Russia) and other geographically close populations like Siberian Mansi, Buryats and other Iranians. New extended HLA haplotypes have been found, such as: A*31:01-B*35:01-DRB1*15:01-DQB1*03:01, A*01:01-B*35:01-DRB1*03:01-DQB1*02:01. Relationships of Turkmen with Kurgan (Gorgan) archaeological mounds, Scythians and Sarmatians are discussed. This study is also useful for a future transplantation Gorgan waiting list, Gorgan HLA and disease epidemiology and HLA pharmacogenomics.

Mixtec Mexican Amerindians: an HLA alleles study for America peopling, pharmacogenomics and transplantation.

HLA-A, -B and -DRB1 alleles have been studied in a Mixtec Mexican Amerindian population by indirect DNA sequencing. HLA relatedness has been tested by comparing results with other Amerindians and worldwide populations; a total of 15,681 chromosomes have been used. Genetic distances between populations, Neighbour Joining (NJ) dendrograms and correspondence analyses have been carried out. Conclusions are: 1) Our Mixtec sample from Oaxaca Coastal Mexican area shows an HLA profile different to that of Oaxaca Central Mountains area showing that genes and languages do not correlate which is inferred both by plane genetic distances and NJ dendrograms and correspondence analyses. 2) Genetic distances and NJ dendrograms join together Mazatecan Mexican Amerindians with our studied Coastal Mixtec group; it fits with the historical relationship between Mixtec and Mazatecans. 3) A*24:02-B*35:14-DRB1*04:11, A*02:01-B*15:15-DRB1*04:11 and A*68:03-B*39:08-DRB1*08:02 extended HLA haplotypes have been "de novo" found in our Mixtec Coastal sample. 4) Shared HLA alleles are found between our Pacific Coast Mixtec Amerindians and Pacific Islanders. 5) These results are useful for establishing a future area transplantation waiting list, for the study of HLA linked diseases epidemiology and for pharmacogenomics in certain drug therapy.

Evaluation of the IL2/IL21, IL2RA and IL2RB genetic variants influence on the endogenous non-anterior uveitis genetic predisposition.

BACKGROUND: Recently, different genetic variants located within the IL2/IL21 genetic region as well as within both IL2RA and IL2RB loci have been associated to multiple autoimmune disorders. We aimed to investigate for the first time the potential influence of the IL2/IL21, IL2RA and IL2RB most associated polymorphisms with autoimmunity on the endogenous non-anterior uveitis genetic predisposition. METHODS: A total of 196 patients with endogenous non-anterior uveitis and 760 healthy controls, all of them from Caucasian population, were included in the current study. The IL2/IL21 (rs2069762, rs6822844 and rs907715), IL2RA (2104286, rs11594656 and rs12722495) and IL2RB (rs743777) genetic variants were genotyped using TaqMan® allelic discrimination assays. RESULTS: A statistically significant difference was found for the rs6822844 (IL2/IL21 region) minor allele frequency in the group of uveitis patients compared with controls (P(-value)=0.02, OR=0.64 CI 95%=0.43-0.94) although the significance was lost after multiple testing correction. Furthermore, no evidence of association with uveitis was detected for the analyzed genetic variants of the IL2RA or IL2RB loci. CONCLUSION: Our results indicate that analyzed IL2/IL21, IL2RA and IL2RB polymorphisms do not seem to play a significant role on the non-anterior uveitis genetic predisposition although further studies are needed in order to clear up the influence of these loci on the non-anterior uveitis susceptibility.

Lack of association between the protein tyrosine phosphatase non-receptor type 22 R263Q and R620W functional genetic variants and endogenous non-anterior uveitis.

OBJECTIVE: Endogenous uveitis is a major cause of visual loss mediated by the immune system. The protein tyrosine phosphatase non-receptor type 22 (PTPN22) gene encodes a lymphoid-specific phosphatase that plays a key role in T-cell receptor (TCR) signaling. Two independent functional missense single nucleotide polymorphisms (SNPs) located within the PTPN22 gene (R263Q and R620W) have been associated with different autoimmune disorders. We aimed to analyze for the first time the influence of these PTPN22 genetic variants on endogenous non-anterior uveitis susceptibility. METHODS: We performed a case-control study of 217 patients with endogenous non-anterior uveitis and 718 healthy controls from a Spanish population. The PTPN22 polymorphisms (rs33996649 and rs2476601) were genotyped using TaqMan allelic discrimination assays. The allele, genotype, carriers, and allelic combination frequencies were compared between cases and controls with χ(2) analysis or Fisher's exact test. RESULTS: Our results showed no influence of the studied SNPs in the global susceptibility analysis (rs33996649: allelic P- value=0.92, odds ratio=0.97, 95% confidence interval=0.54-1.75; rs2476601: allelic P- value=0.86, odds ratio=1.04, 95% confidence interval=0.68-1.59). Similarly, the allelic combination analysis did not provide additional information. CONCLUSIONS: Our results suggest that the studied polymorphisms of the PTPN22 gene do not play an important role in the pathophysiology of endogenous non-anterior uveitis.

HLA in Georgians (Caucasus) and their relationship with Eastern Mediterraneans.

HLA-A, -B, -DQB1, and -DRB1 typing has been performed in a sample of Georgian population (South Caucasus). Allele frequencies, neighbour joining and correspondence relatedness analyses and extended HLA haplotypes have been obtained with comparison with other Middle East and Mediterranean populations. Our Georgian sample tends to be genetically related in these analyses with Eastern Mediterraneans and Middle East people. This is important for future regional transplant programs, and Georgian HLA and disease epidemiology and pharmacogenomics.

Major histocompatibility complex complement (MHC) Bf alleles show trans species evolution between man and chimpanzee.

HLA and disease studies by using single allele statistics have been fruitless during the last 40 years for explaining association pathogenesis of the associated diseases.Other approaches are necessary to untangle this puzzle. We aim to revisit complement alleleism in humans and primates for both studying MHC and disease association to complotypes and extended MHC haplotypes in order to also explain the positive directional selection of maintaining immune response genes (complement, MHC adaptive and MHC non-specific genes) that keeps these three type of genes together in a short chromosome stretch (MHC) for million years. These genes may be linked to conjointly avoid microbes attack and autoimmunity. In the present paper, it is obtained a new Bf chimpanzee allele, provisionaly named Patr-Bf*A:01,that differs from other Bf alleles by having CTG at eleventh codon of exon 2 in order to start the newly suggested methodology and explain functional and evolutionary MHC obscure aspects. Exons 1 to 6 of Ba fragment of Bf gene were obtained from chimpanzee. This new chimpanzee Factor B allele (Patr-Bf*A:01) is to be identical to a infrequent human Bf allele (SNP rs641153); it stresses the strong evolutive pressure upon certain alleles that are trans specific. It also may apply to MHC extended haplotipes which may conjointly act to start an adequate immune response. It is the first time that a complement MHC class III allele is described to undergo trans species evolution,in contrast to class I and class II alleles which had already been reported . Allelism of complement factors are again proposed for studying MHC complement genes, complotypes, and extended MHC haplotypes which may be more informative that single MHC marker studies.

HLA study in Mexico Nahua/Aztec Amerindians: Close relatedness to the ancient Central America ethnic groups.

Nahua population (also named Aztec or Mexica) was studied for HLA class II genes in a Mexican rural city (Santo Domingo Ocotitlan, Morelos State) belonging to the nowadays Náhuatl speaking areas in Mexico. The most frequent HLA class II alleles were typical Amerindian (HLA-DRB1*04:07, DQB1*03:01 DRB1*04:03 or DRB1*04:04) and also were some calculated extended haplotypes (HLA-DRB1*04:07-DQB1*03:02,DRB1*08:02-DQB1*04:02, or DRB1*10:01-DQB1*05:01 among others). When using HLA-DRB1 Neís genetic distances, our isolated Nahua population was found to be close to other Central America Amerindians like the ancient-established Mayans or Mixe. This may suggest that Nahuas origin was also from Central America. It contrasts to legend that assumes they came from the North, and they built the Aztec Empire after submitting Central America neighbouring ethnic groups before 1519 CE when Spaniards led by Hernán Cortés arrived to Mexico.

HLA alleles and haplotypes in Sudanese population and their relationship with Mediterraneans.

The contribution of migrated people from once green Sahara (about 10,000-6000 years BC) towards Mediterranean area had probably a double effect: both genetic and cultural connections have been described between Western Europe and North Africa. Sudanese populations from different ethnicities have been studied for HLA-A, -B, -DRB1 and -DQB1 antigens by a standard microlymphotoxicity method. Results found show that Nubians are genetically related with African Sub-Saharan populations and distant from other Sudanese tribes, who are closer to Mediterranean populations than to Sub-Saharan ones. This is concordant with other authors and meta-analysis data. Our present work is, to our knowledge, the first and only one HLA research that studies Sudanese people according to different Sudan ethnic groups: samples were collected before Sudan partition between North and South. A prehistoric genetic and peoples exchange between Africa and the Mediterranean basin may be observed and is supported with the results obtained in this Sudanese HLA study. However, demic diffusion model of agriculture and other anthropological traits from Middle East to West Europe/Maghreb do not exist: a more detailed Sahel and North African countries ancient and recent admixture studies are also being carried out which may clearer explain pastoralists/agriculture innovations origins in Eurafrican Mediterranean and Atlantic façade.

Genetic signature detected in T cell receptors from patients with severe COVID-19.

Characterization of host genetic factors contributing to COVID-19 severity promises advances on drug discovery to fight the disease. Most genetic analyses to date have identified genome-wide significant associations involving loss-of-function variants for immune response pathways. Despite accumulating evidence supporting a role for T cells in COVID-19 severity, no definitive genetic markers have been found to support an involvement of T cell responses. We analyzed 205 whole exomes from both a well-characterized cohort of hospitalized severe COVID-19 patients and controls. Significantly enriched high impact alleles were found for 25 variants within the T cell receptor beta (TRB) locus on chromosome 7. Although most of these alleles were found in heterozygosis, at least three or more in TRBV6-5, TRBV7-3, TRBV7-6, TRBV7-7, and TRBV10-1 suggested a possible TRB loss of function via compound heterozygosis. This loss-of-function in TRB genes supports suboptimal or dysfunctional T cell responses as a major contributor to severe COVID-19 pathogenesis.

HLA-G: Too Much or Too Little? Role in Cancer and Autoimmune Disease.

HLA-G is a non-classical HLA class I molecule with immunomodulatory properties. It was initially described at the maternal-fetal interface, and it was later found that this molecule was constitutively expressed on certain immuneprivileged tissues, such as cornea, endothelial and erythroid precursors, and thymus. The immunosuppressive effect of HLA-G is exerted through the interaction with its cognate receptors, expressed on immunocompetent cells, like ILT2, expressed on NK, B, T cells and APCs; ILT4, on APCs; KIR, found on the surface of NK cells; and finally, the co-receptor CD8. Because of these immunomodulatory functions, HLA-G has been involved in several processes, amongst which organ transplantation, viral infections, cancer progression, and autoimmunity. HLA-G neo-expression on tumors has been recently described in several types of malignancies. In fact, tumor progression is tightly linked to the presence of the molecule, as it exerts its tolerogenic function, inhibiting the cells of the immune system and favoring tumor escape. Several polymorphisms in the 3'UTR region condition changes in HLA-G expression (14bp and +3142C/G, among others), which have been associated with both the development and outcome of patients with different tumor types. Also, in recent years, several studies have shown that HLA-G plays an important role in the control of autoimmune diseases. The ability of HLA-G to limit the progression of these diseases has been confirmed and, in fact, levels of the molecule and several of its polymorphisms have been associated with increased susceptibility to the development of autoimmune diseases, as well as increased disease severity. Thus, modulating HLA-G expression in target tissues of oncology patients or patients with autoimmune diseases may be potential therapeutic approaches to treat these pathological conditions.

A Reliable and Standardizable Differential PCR and qPCR Methodology Assesses HER2 Gene Amplification in Gastric Cancer.

We have applied two PCR techniques, differential PCR (diffPCR) and qPCR for the identification of HER2 gene amplifications in genomic DNA of tumor and distal gastric samples from patients with gastric cancer. The diffPCR technique consists of the simultaneous amplification of the HER2 gene and a housekeeping gene by conventional PCR and the densitometric analysis of the bands obtained. We established a cut-off point based on the mean and standard deviation analyzing the DNA of 30 gastric tissues from patients undergoing non-cancer gastrectomy. diffPCR and qPCR yielded consistent results. HER2-overexpression was detected in 25% of patients and was further confirmed by immunohistochemistry and immunofluorescence. The approaches herein described may serve as complementary and reliable methods to assess HER2 amplification.

HLA-G 3'UTR Polymorphisms Are Linked to Susceptibility and Survival in Spanish Gastric Adenocarcinoma Patients.

HLA-G is a non-classical class I HLA molecule that induces tolerance by acting on receptors of both innate and adaptive immune cells. When overexpressed in tumors, limits surveillance by the immune system. The HLA-G gene shows several polymorphisms involved in mRNA and protein levels. We decided to study the implication of two polymorphisms (rs371194629; 14bp INS/DEL and rs1063320; +3142 C/G) in paired tissue samples (tumoral and non-tumoral) from 107 Spanish patients with gastric adenocarcinoma and 58 healthy control individuals, to assess the possible association of the HLA-G gene with gastric adenocarcinoma susceptibility, disease progression and survival. The presence of somatic mutations involving these polymorphisms was also analyzed. The frequency of the 14bp DEL allele was increased in patients (70.0%) compared to controls (57.0%, p=0.025). In addition, the haplotype formed by the combination of the 14bp DEL/+3142 C variants is also increased in patients (54.1% vs 44.4%, p=0.034, OR=1.74 CI95% 1.05-2.89). Kaplan-Meier analysis revealed that 14bp DEL/DEL patients showed lower 5-year life-expectancy than INS/DEL or INS/INS (p=0.041). Adjusting for TNM staging (Cox regression analysis) disclosed a significant difference in death risk (p=0.03) with an expected hazard 2.6 times higher. Finally, no somatic mutations were found when comparing these polymorphisms in tumoral vs non-tumoral tissues, which indicates that this is a preexisting condition in patients and not a de novo, tumor-restricted, event. In conclusion, the variants predominant in patients were those increasing HLA-G mRNA stability and HLA-G expression, clearly involving this molecule in gastric adenocarcinoma susceptibility, disease progression and survival and making it a potential target for immunotherapeutic approaches.

An Experimental DUAL Model of Advanced Liver Damage.

Individuals exhibiting an intermediate alcohol drinking pattern in conjunction with signs of metabolic risk present clinical features of both alcohol-associated and metabolic-associated fatty liver diseases. However, such combination remains an unexplored area of great interest, given the increasing number of patients affected. In the present study, we aimed to develop a preclinical DUAL (alcohol-associated liver disease plus metabolic-associated fatty liver disease) model in mice. C57BL/6 mice received 10% vol/vol alcohol in sweetened drinking water in combination with a Western diet for 10, 23, and 52 weeks (DUAL model). Animals fed with DUAL diet elicited a significant increase in body mass index accompanied by a pronounced hypertrophy of adipocytes, hypercholesterolemia, and hyperglycemia. Significant liver damage was characterized by elevated plasma alanine aminotransferase and lactate dehydrogenase levels, extensive hepatomegaly, hepatocyte enlargement, ballooning, steatosis, hepatic cell death, and compensatory proliferation. Notably, DUAL animals developed lobular inflammation and advanced hepatic fibrosis. Sequentially, bridging cirrhotic changes were frequently observed after 12 months. Bulk RNA-sequencing analysis indicated that dysregulated molecular pathways in DUAL mice were similar to those of patients with steatohepatitis. Conclusion: Our DUAL model is characterized by obesity, glucose intolerance, liver damage, prominent steatohepatitis and fibrosis, as well as inflammation and fibrosis in white adipose tissue. Altogether, the DUAL model mimics all histological, metabolic, and transcriptomic gene signatures of human advanced steatohepatitis, and therefore serves as a preclinical tool for the development of therapeutic targets.