Celiac disease diagnosis: transglutaminase, duodenal biopsy and genetic tests correlations.

Introduction: Celiac disease (CD) is an autoimmune enteropathy triggered by gluten ingestion in genetically susceptible individuals. The haplotypes HLA-DQ2 and DQ8, transglutaminase (TGA) antibodies, and biopsy findings are the main tests performed in the evaluation and CD diagnosis. The objective was to establish possible correlations between transglutaminase levels, genetic markers tests, and qualitative intestinal biopsy findings (modified Marsh classification) at the diagnosis. Methods: A retrospective cohort study. The selection criteria were confirmed CD cases with genetic tests performed. Statistical analysis was done mainly through One-way ANOVA, Kendall's correlation coefficient (T), and linear regression. Results: The study included 112 patients, with a mean age of 6 ± 4 years. All cases were tested to HLA-DQ2, and it was positive in 93%. HLA-DQ8 was tested in 73% of cases and it was positive in 61%. The percentage of negative genetic markers (DQ2/DQ8) was 4.5% for patients tested to both haplotypes. A comparison of DQ2/DQ8 (positive and negative) with clinical findings and tests performed did not identify any differences for most of the parameters analyzed. Cases of type I diabetes presented significant negative expression for DQ2(-); p = 0.05 and positive expression for DQ8(+); p = 0.023. The TGA antibody levels ranged from 18 to 36,745 U/ml. An inverse correlation was found between age and TGA-L level (p = 0.043). In 23% of the cases, the TGA levels were greater than 1,000 U/ml and presented a moderate positive correlation with the atrophy biopsy profile (T = 0.245). Patients with an atrophic biopsy profile (Marsh III) had a moderate positive correlation with growth failure (T = 0.218) but a negative correlation with constipation (T = -0.277). Conclusion: In terms of diagnosis tests for CD, transglutaminase levels and age presented an inverse correlation, with the level decreasing as age increased. A moderately positive correlation was found between mean transglutaminase with intestinal atrophy and growth retardation. The genetic test DQ2 was positive for 93% and negative genetic markers (DQ2/DQ8) represented 4.5% of cases studied.

Almond protein as Pickering emulsion stabilizer: Impact of microgel fabrication method and pH on emulsion stability.

We evaluated the ability of almond proteins to produce Pickering emulsions (EM) stabilized by microgels (MG) fabricated by three different methods (heat treatment-HT, crosslinking with transglutaminase-TG or calcium-CA), at two pH levels (pH 3 or 7). Compared to pH 7, acidic pH significantly denatured almond proteins (ellipticity ∼0 mdeg), decreased absolute zeta potential values (10.5 to 18.6 mV at pH 3 and - 24.6 to -32.6 mV at pH 7), and free thiol content (114.64-131.60 µmol SH/g protein at pH 3 and 129.46-148.17 µmol SH/g protein at pH 7 - except in CA-crosslinked microgels, p > 0.05). These changes led to larger microgel sizes (D3,2pH3: 26.3-39.5 µm vs. D3,2pH7: 5.9-9.0 µm) with lower polydispersity (SpanpH3: ∼ 1.94 vs. SpanpH7: 2.32, excluding CA-based samples). Consequently, the Turbiscan Stability Index (TSI) was higher in acidic conditions for all emulsions, except for the calcium-containing formulation (EM_CApH3), emphasizing the critical role of calcium binding in maintaining emulsion stability in acidic environments. Microgels prepared via the traditional heat treatment method produced emulsions with intermediate stability (TSI ranging from 3.4 % to 5.1 % at 28 days of storage). Conversely, TG-crosslinked microgels led to unstable emulsions at pH 3, likely due to the lowest zeta potential (+4.2 mV), whereas at pH 7, the greatest stability was attributed to bridging flocculation that created a stable gel-like structure. Indeed, emulsions with lower TSI (EM_CApH3 = 1.8 %, EM_CApH7 = 2.3 % and EM_TGpH7 = 1.0 %, at 28 days of storage) also exhibited higher elastic modulus (G') over frequency sweep, indicating that the strong elastic network was relevant for emulsion stability (up to 28 days). This study, for the first time, demonstrated the production of stable almond-based Pickering emulsions, with properties modulated by the pH and method used to fabricate the microgels.

Fish Fillet Analogue Using Formulation Based on Mushroom (Pleurotus ostreatus) and Enzymatic Treatment: Texture, Sensory, Aromatic Profile and Physicochemical Characterization.

The growing demand for alternative sources of non-animal proteins has stimulated research in this area. Mushrooms show potential in the innovation of plant-based food products. In this study, the aim was to develop prototype fish fillets analogues from Pleurotus ostreatus mushrooms applying enzymatic treatment (ß-glucanase and transglutaminase-TG). A Plackett-Burman 20 experimental design was used to optimize forty variables. Oat flour (OF) exerted a positive effect on the hardness and gumminess texture parameters but a negative effect on cohesiveness and resilience. Soy protein isolate (SPI) exhibited a positive effect on elasticity, gumminess and chewiness, while acacia gum had a negative effect on elasticity, cohesiveness and resilience. After sensory analysis the assay with 1% cassava starch, 5% OF, 5% SPI, 0.1% transglutaminase (240 min/5 °C), 1% coconut oil, 1% soybean oil, 0.2% sodium tripolyphosphate, 0.6% ß-glucanase (80 °C/10 min) and without ß-glucanase inactivation was found to exhibit greater similarity to fish fillet. The classes hydrocarbons, alcohols and aldehydes are the predominant ones in aromatic profile analysis by chromatography and electronic nose. It is concluded that a mushroom-based analogue of fish fillet can be prepared using enzymatic treatment with TG.

Evaluation of HLA-DQ2 and HLA-DQ8 haplotypes in patients with endometriosis, A case-control study.

OBJECTIVE: To evaluate the relationship between genetic haplotypes associated with celiac disease (Human Leucocyte Antigen [HLA] DQ2 and DQ8) with the diagnosis, clinical presentation, and location of endometriosis in Brazilian women. METHOD: A retrospective cross-sectional study, was conducted in a Tertiary hospital. PATIENTS: Women aged 18-50 years who underwent HLA-DQ2 and HLA-DQ8 haplotype analysis. INTERVENTION: The patients were divided into endometriosis and control groups and evaluated for symptoms; endometriosis location, American Society for Reproductive Medicine (ASRM) stage, and the presence of anti-tissue transglutaminase IgA (anti-TgA), HLA-DQ2, and HLA-DQ8 markers. RESULTS: A total of 434 consecutive patients with (n = 315) and without (n = 119) endometriosis were included. Pain and infertility were more frequent in the endometriosis group than in the control group. The presence of HLA-DQ2, HLA-DQ8, and anti-TgA was similar between both groups. The presence of HLA-DQ2 and HLA-DQ8 markers did not differ based on age, pain symptoms, ASRM stage, or endometriosis location. CONCLUSION: Although there are similarities in inflammatory markers and pathophysiology between celiac disease and endometriosis, this study found no significant associations in the presence of HLA-DQ2 or HLA-DQ8 haplotypes and endometriosis.

Addition of transglutaminase enzyme in camel milk yoghurt to increase its sensorial aspects / Adição de enzima transglutaminase em iogurte de leite de camela para aumento de aspectos sensoriais

Camel milk was obtained from A-block UVAS Ravi Campus Pattoki. After pasteurization at 72 °C (15 sec) it was cooled to 42 °C, then glutathione treated transglutaminase enzyme was added with the concentration of 0.5 g/300 mL, 1 g/300 mL, 1.5 g/300 mL, 2 g/300 mL while control sample with the addition of 1.5 g/300 mL gelatin. Then inoculation of milk was done with standard cultures of Yoghurt Lactobacillus delbrueckii subsp bulgaricus and Streptococcus thermophilus at the rate of 2% for 3-4 hours at 42 °C. Samples were stored at 4 °C and were analyzed on 1st day and 28th day of storage. In our findings, there was slight increase in sensorial properties of all the samples. It was also observed that syneresis was reduced with the increase of enzyme quantity.

O leite de camelo foi obtido do bloco B Uvas Ravi Campus Pattoki. Após a pasteurização a 72 °C (15 s), foi resfriado a 42 °C, posteriomente, à enzima transglutaminase tratada com glutationa foi adicionada com a concentração de 0,5 g/300 mL, 1 g/300 mL, 1,5 g/300 mL, 2 g/300 mL enquanto a amostra controle com a adição de 1,5 g/300 mL de gelatina. Em seguida, a inoculação do leite foi feita com culturas padrão de Iogurte Lactobacillus delbrueckii subsp bulgaricus e Streptococcus thermophilus com taxa de 2% durante 3-4 horas a 42 °C. As amostras foram armazenadas a 4 °C e analisadas no 1º dia e no 28º dia de armazenamento. Em nossos achados, houve leve aumento nas propriedades sensoriais de todas as amostras. Observou-se também que a sinérese foi reduzida com o aumento da quantidade da enzima.

Evaluation of HLA-DQ2 and HLA-DQ8 haplotypes in patients with endometriosis, A case-control study

Abstract Objective To evaluate the relationship between genetic haplotypes associated with celiac disease (Human Leucocyte Antigen [HLA] DQ2 and DQ8) with the diagnosis, clinical presentation, and location of endometriosis in Brazilian women. Method A retrospective cross-sectional study, was conducted in a Tertiary hospital. Patients Women aged 18-50 years who underwent HLA-DQ2 and HLA-DQ8 haplotype analysis. Intervention The patients were divided into endometriosis and control groups and evaluated for symptoms; endometriosis location, American Society for Reproductive Medicine (ASRM) stage, and the presence of anti-tissue transglutaminase IgA (anti-TgA), HLA-DQ2, and HLA-DQ8 markers. Results A total of 434 consecutive patients with (n = 315) and without (n = 119) endometriosis were included. Pain and infertility were more frequent in the endometriosis group than in the control group. The presence of HLA-DQ2, HLA-DQ8, and anti-TgA was similar between both groups. The presence of HLA-DQ2 and HLA-DQ8 markers did not differ based on age, pain symptoms, ASRM stage, or endometriosis location. Conclusion Although there are similarities in inflammatory markers and pathophysiology between celiac disease and endometriosis, this study found no significant associations in the presence of HLA-DQ2 or HLA-DQ8 haplotypes and endometriosis.

Prevalence of IgA anti-tissue transglutaminase antibody in a cohort of iranians patients with inflammatory bowel disease

Background and Aims: Some studies have reported the coexistence of inflammatory bowel disease (IBD) and celiac disease (CD). However, the prevalence of anti-tissue transglutaminase antibodies (IgA and IgG) and their screening value in patients with IBD is not yet clear. This study aimed to assess the prevalence of IgA anti-tTG and its potential correlation with disease status in patients with IBD. Materials and Methods: This cross-sectional study was conducted on 110 patients with confirmed IBD diagnosis at Ghaem Hospital, Mashhad, Iran. For each patient, all demographic and clinical data including age, extra intestinal manifestations, underlying diseases, types of diseases, and surgical history were collected. IgA anti-tissue transglutaminase titers were assessed by enzyme-linked immunosorbent assay. Results: None of the patients with IBD were positive for IgA anti-tTG antibodies, with a mean titer of 3.31 ± 1.3 AU/mL. Also, the mean titers were not associated with age, gender and various disease clinical features including the disease history, underlying disease, diagnosis type, extraintestinal manifestations, and surgery history. Conclusion: No significant prevalence pattern of IgA anti-tTG antibody was observed in patients with IBD. Accordingly, serological screening for CeD is not recommended in IBD patients, unless in a relevant clinical CeD suspicion. (AU)

Celiac disease seroprevalence in subjects with dyspeptic symptoms. A study on a Mexican population.

INTRODUCTION AND AIMS: Celiac disease (CD) is an autoimmune enteropathy that develops in genetically susceptible individuals. The typical gastrointestinal manifestation is diarrhea but symptoms of dyspepsia, such as epigastric pain, nausea, or satiety, can sometimes appear. Previous studies have reported that the prevalence of CD in patients with dyspepsia can be as high as 7%. The aim of the present study was to evaluate CD seroprevalence in subjects with dyspeptic symptoms and a control group in a Mexican population. MATERIAL AND METHODS: A case-control study was conducted on blood donors that answered the PAGI-SYM questionnaire for dyspepsia and in whom IgA antibodies to tissue transglutaminase 2 (IgA anti-tTG2) and IgG antibodies to deamidated gliadin peptide (IgG anti-DGP) were determined. CD seroprevalence in subjects with dyspeptic symptoms and in asymptomatic subjects was compared. RESULTS: A total of 427 subjects (76.3% men), with a mean patient age of 34 years (range of 18-65 years) were included. Of those participants, 87 (20.3%) had symptoms of dyspepsia (group A) and 340 (79.6%) were asymptomatic (group B). Antibodies were positive in one (1.15%) of the group A subjects (1/87, 95% CI 0.2-6 %), whereas they were positive in 4 (1.18%) of the group B subjects (4/340, 95% CI 0.4-2.9%, p = 0.59). CONCLUSIONS: CD seroprevalence in the study population with dyspeptic symptoms (1%) was not different from that of the control population. Thus, CD screening in Mexican patients with dyspepsia is not justified.

TG2 promotes amyloid beta aggregates: Impact on ER-mitochondria crosstalk, calcium homeostasis and synaptic function in Alzheimer's disease.

Alzheimer's Disease (AD) is a neurodegenerative disorder characterized by cognitive impairment that increasingly affects the elderly. AD's main features have been related to cellular and molecular events, including the aberrant aggregation of the amyloid beta peptide (Aß), Ca2+ dyshomeostasis, and increased mitochondria-associated membranes (MAMs). Transglutaminase type 2 (TG2) is a ubiquitous enzyme whose primary role is the Ca2+-dependent proteins transamidation, including the Aß peptide. TG2 activity has been closely related to cellular damage and death. We detected increased TG2 levels in neuronal cells treated with Aß oligomers (AßOs) and hippocampal slices from J20 mice using cellular and molecular approaches. In this work, we characterized the capacity of TG2 to interact and promote Aß toxic aggregates (AßTG2). AßTG2 induced an acute increase in intracellular Ca2+, miniature currents, and hiperexcitability, consistent with an increased mitochondrial Ca2+ overload, IP3R-VDAC tethering, and mitochondria-endoplasmic reticulum contacts (MERCs). AßTG2 also decreased neuronal viability and excitatory postsynaptic currents, reinforcing the idea of synaptic failure associated with MAMs dysregulation mediated by TG2. Z-DON treatment, TG2 inhibitor, reduced calcium overload, mitochondrial membrane potential loss, and synaptic failure, indicating an involvement of TG2 in a toxic cycle which increases Aß aggregation, Ca2+ overload, and MAMs upregulation. These data provide novel information regarding the role TG2 plays in synaptic function and contribute additional evidence to support the further development of TG2 inhibitors as a disease-modifying strategy for AD.

Mechanism study of the gel-forming ability of heat-induced gel from Peruvian hake (Merluccius gayi peruanus) surimi.

The commercial value of Peruvian hake (Merluccius gayi peruanus) meat is low because of its soft texture. This study investigated the major factor contributing to the gel-forming ability of Peruvian hake surimi by comparing the effects of endogenous protease activity and parasitic infection. Heat-induced gels could not be obtained at 50 °C-90 °C. Surimi with severe parasitic infection showed a stronger gel-forming ability. The endogenous protease activities were the main factor influencing the Peruvian hake meat proteolysis and contributed to the low gel-forming ability, rather than parasitic infection. Specifically, endogenous cysteine proteases played an essential role in protein degradation and low gel-forming ability. Moreover, endogenous transglutaminase was also shown to be involved in the gel-forming ability upon heating at 40 °C. These results suggested that Peruvian hake meat could be used as a raw material of frozen surimi for fish gel by inhibiting the activity of endogenous proteases.

Enzymatic Crosslinked Hydrogels of Gelatin and Poly (Vinyl Alcohol) Loaded with Probiotic Bacteria as Oral Delivery System.

Probiotic bacteria are widely used to prepare pharmaceutical products and functional foods because they promote and sustain health. Nonetheless, probiotic viability is prone to decrease under gastrointestinal conditions. In this investigation, Lactiplantibacillus plantarum spp. CM-CNRG TB98 was entrapped in a gelatin−poly (vinyl alcohol) (Gel−PVA) hydrogel which was prepared by a "green" route using microbial transglutaminase (mTGase), which acts as a crosslinking agent. The hydrogel was fully characterized and its ability to entrap and protect L. plantarum from the lyophilization process and under simulated gastric and intestine conditions was explored. The Gel−PVA hydrogel showed a high probiotic loading efficiency (>90%) and survivability from the lyophilization process (91%) of the total bacteria entrapped. Under gastric conditions, no disintegration of the hydrogel was observed, keeping L. plantarum protected with a survival rate of >94%. While in the intestinal fluid the hydrogel is completely dissolved, helping to release probiotics. A Gel−PVA hydrogel is suitable for a probiotic oral administration system due to its physicochemical properties, lack of cytotoxicity, and the protection it offers L. plantarum under gastric conditions.

Anticuerpos antitransglutaminasa tisular en pacientes con síntomas gastrointestinales / Anti-tissue transglutaminase antibodies in patients with gastrointestinal symptoms

Introducción: La enfermedad celiaca es una enteropatía mediada por la respuesta inmune, que ha sido crecientemente reconocida como una enfermedad común, que afecta tanto a la población infantil, como a la adulta. La serología es un componente clave de la detección y diagnóstico de la celiaquía. Objetivo: Evaluar la utilidad diagnóstica de los anticuerpos antitransglutaminasa tisular en individuos con síntomas gastrointestinales crónicos. Métodos: En un estudio de corte se determinaron los anticuerpos anti-transglutaminasa tisular IgA/G en 87 pacientes adultos y pediátricos con indicación médica de anticuerpos de celiaquía. Los anti- transglutaminasa tisular IgA/G se realizaron por el ensayo inmunoadsorbente ligado a enzima y por el ensayo multiplex de inmunoblot. Se aplicó la prueba U de Mann-Whitney y se calculó el coeficiente de concordancia kappa. Resultados: La seroprevalencia de los anti-transglutaminasa tisular IgG/IgA resultó de 8,05 por ciento (7/87) por el ensayo inmunoenzimático. Los resultados cualitativos del ensayo inmunoenzimático y del inmunoblot para los anti- transglutaminasa tisular fueron concordantes con un coeficiente kappa de 0,407 (p=0,004). La distribución de la concentración de los anticuerpos anti-TGt IgA/G obtenidos por el ensayo inmunoenzimático respecto a los resultados negativos y positivos del inmunoblot no fue significativa (p=0,08). Los pacientes con presencia de anti-transglutaminasa tisular IgA/G por el ensayo inmunoenzimático obtuvieron el diagnóstico definitivo de enfermedad celiaca confirmado por biopsia duodenal. Conclusiones: Se confirmó la utilidad de la detección de los anticuerpos anti-transglutaminasa tisular IgA/G por el ensayo inmunoenzimático como primer paso diagnóstico de la enfermedad celíaca en pacientes con síntomas gastrointestinales(AU)

Introduction: Celiac disease is an immune-mediated enteropathy that has been increasingly recognized as a common disease, affecting both the pediatric and adult population. Serology is a key component of the detection and diagnosis of celiac disease. Objective: To evaluate the diagnostic usefulness of anti-tissue transglutaminase antibodies in individuals with chronic gastrointestinal symptoms. Methods: In a cutoff study, anti-tissue transglutaminase IgA/G antibodies were determined in 87 adult and pediatric patients with medical indication for celiac disease antibodies. Anti-tissue transglutaminase IgA/G was performed by enzyme-linked immunoadsorbent assay and multiplex immunoblot assay. Mann-Whitney U test was applied and kappa correspondence coefficient was calculated. Results: The seroprevalence of anti-tissue transglutaminase IgG/IgA was 8.05 percent (7/87) by enzyme-linked immunosorbent assay. The qualitative results of the enzyme-linked immunosorbent assay and immunoblot for anti-tissue transglutaminase were consistent with a kappa coefficient of 0.407 (p=0.004). The distribution of the concentration of anti-TGt IgA/G antibodies obtained by enzyme-linked immunosorbent assay with respect to negative and positive immunoblot results was not significant (p=0.08). Patients with presence of anti-tissue transglutaminase IgA/G by enzyme-linked immunosorbent assay obtained the definitive diagnosis of celiac disease confirmed by duodenal biopsy. Conclusions: The usefulness of detection of anti-tissue transglutaminase IgA/G antibodies by enzyme-linked immunosorbent assay as a first diagnostic step of celiac disease in patients with gastrointestinal symptoms was confirmed(AU)

HLA DQ2/DQ8 haplotypes and anti-transglutaminase antibodies as celiac disease markers in a pediatric population with type 1 diabetes mellitus.

Objective: Evaluate the celiac disease (CD) markers, within the scope of its screening, in a pediatric population with diagnosis of type 1 diabetes (T1D) at Hospital de Braga (HB) and determine the prevalence of CD in the sample. Reflect on CD screening algorithm applied in this pediatric population. Methods: Retrospective observational study with 94 patients diagnosed with T1D at age 10 years or younger, followed up at the HB Outpatient Diabetology Consultation, including those referred from other hospitals. Record of clinical information, IgA anti-transglutaminase and anti-endomysium and HLA DQ2/DQ8 haplotypes. Results: We obtained positive serological test for CD in 4 patients. This test had 100% sensitivity and specificity. The prevalence of CD was 4.3% (n = 4). Positive HLA screening in 84.6% of patients, with both sensitivity and negative predictive value of 100% and specificity of 16.67%. Diagnosis of CD was made on average 3.40 ± 3.32 years after the diagnosis of TD1. All cases of CD registered non-gastrointestinal manifestations, none had gastrointestinal symptoms. Conclusion: This study proved that there is a higher prevalence of CD in pediatric population with TD1, when compared to general population, and clarified the importance of CD screening. Furthermore, it was observed that serological screening for CD antibodies is an excellent screening test and HLA typing, although not the most suitable first line test, can be useful in excluding the possibility of patients with T1D developing CD.

HLA DQ2/DQ8 haplotypes and anti-transglutaminase antibodies as celiac disease markers in a pediatric population with type 1 diabetes mellitus

ABSTRACT Objectives: Evaluate the celiac disease (CD) markers, within the scope of its screening, in a pediatric population with diagnosis of type 1 diabetes (T1D) at Hospital de Braga (HB) and determine the prevalence of CD in the sample. Reflect on CD screening algorithm applied in this pediatric population. Subjects and methods: Retrospective observational study with 94 patients diagnosed with T1D at age 10 years or younger, followed up at the HB Outpatient Diabetology Consultation, including those referred from other hospitals. Record of clinical information, IgA anti-transglutaminase and anti-endomysium and HLA DQ2/DQ8 haplotypes. Results: We obtained positive serological test for CD in 4 patients. This test had 100% sensitivity and specificity. The prevalence of CD was 4.3% (n = 4). Positive HLA screening in 84.6% of patients, with both sensitivity and negative predictive value of 100% and specificity of 16.67%. Diagnosis of CD was made on average 3.40 ± 3.32 years after the diagnosis of TD1. All cases of CD registered non-gastrointestinal manifestations, none had gastrointestinal symptoms. Conclusion: This study proved that there is a higher prevalence of CD in pediatric population with TD1, when compared to general population, and clarified the importance of CD screening. Furthermore, it was observed that serological screening for CD antibodies is an excellent screening test and HLA typing, although not the most suitable first line test, can be useful in excluding the possibility of patients with T1D developing CD.

Hypoxia and inflammation conditions differentially affect the expression of tissue transglutaminase spliced variants and functional properties of extravillous trophoblast cells.

PROBLEM: Persistent hypoxia and inflammation beyond early pregnancy are involved in a bad outcome because of defective trophoblast invasiveness. Tissue transglutaminase (TG2) coregulates several cell functions. An aberrant expression and/or transamidation activity could contribute to placental dysfunction. METHOD OF STUDY: The first-trimester trophoblast cell line (Swan-71) was used to study TG2 expression and cell functions in the absence or presence of inflammatory cytokines (TNF-α, IL-1ß) or chemical hypoxia (CoCl2 ). We analyzed The concentration of cytokines in the supernatant by ELISA; Cell migration by scratch assay; NF-κB activation by detection of nuclear p65 by immunofluorescence or flow cytometry using a Swan-71 NF-κB-hrGFP reporter cell line. Tissue transglutaminase expression was analyzed by immunoblot and confocal microscopy. Expression of spliced mRNA variants of tissue transglutaminase was analyzed by RT-PCR. Transamidation activity was assessed by flow cytometry using 5-(biotinamido)-pentylamine substrate. RESULTS: Chemical hypoxia and TGase inhibition, but not inflammatory stimuli, decreased Swan-71 migration. IL-6 production was also decreased by chemical hypoxia, but increased by inflammation. Intracellular TGase activity was increased by all stimuli, but NF-κB activation was observed only in the presence of proinflammatory cytokines. TG2 expression was decreased by CoCl2 and TNF-α. Translocation of TG2 and p65 to nuclei was observed only with TNF-α, without colocalization. Differential relative expression of spliced variants of mRNA was observed between CoCl2 and inflammatory stimuli. CONCLUSION: The observed decrease in total TG2 expression and relative increase in short variants under hypoxia conditions could contribute to impaired trophoblast invasion and impact on pregnancy outcome.

Histological changes and transglutaminase 2 expression in the oviduct of advanced pregnant cows.

The oviduct is a dynamic organ that has not been assigned specific functions during advanced pregnancy. However, since changes in the oviductal epithelium during the estrous cycle are attributed mainly to variations in estradiol (E2) levels, and E2 levels increase along pregnancy, we hypothesized that advanced pregnant cows should present changes in the oviductal epithelium. In advanced pregnant cows, the oviducts showed higher leaf-like folds and lower mucosa width and epithelium height than those of cycling animals. Also, PAS-positive apical protrusions and TUNEL-positive extruded cytoplasmic material were observed in advanced pregnant cows. Oviductal fluid from advanced pregnant cows showed lower protein concentration than that from cycling cows. Transglutaminase 2 (TG2) was detected exclusively in oviductal fluid of pregnant cows but not in cells from any stage, whereas its mRNA was detected in different amounts in cells from all stages. This protein was identified by LC/MS-MS and its identity was corroborated by Western blot. The observations in histology of the epithelium and the presence of TG2 in oviductal fluid correlate with high levels of E2 in serum. In conclusion, important histological changes in the oviductal epithelium and secretion of TG2 to the oviductal fluid appear to be triggered by the high E2 levels exclusive of advanced pregnancy.

Improving of Gelling Capacity of Cooked Crab Meat Proteins.

Cooked crab meat subjected to a cutting process can aggregate again, forming weak gels. The objective of this work was to determine the effect of two mixing methods, combined with the addition of the microbial enzyme TGase (MTGase) on the mechanical and functional properties of gels from washed or unwashed blue crab (Callinectes sapidus) meat. Live crabs were obtained from Laguna Madre, Tamaulipas, Mexico, and cooked at 120°C for 20 min before hand-picking the meat from the shell. Cooked meat was processed by mixing and cut at temperatures of 25 or 60°C, without (control) or 0.5% of MTGase. Then cooked at 90°C for 15 min. Changes in texture profile analysis, percentage of extractable water, and color were evaluated. The mixing method at 60°C allowed increasing the textural properties of the gels, and the addition of MTGase significantly improved the mechanical properties. The results allowed stablishing a viable technique to obtain restructured gels from cooked crab meat with no need to extract the soluble compounds responsible for their distinctive odor and taste which often affect the mechanical properties.

Expression of Bacillus amyloliquefaciens transglutaminase in recombinant E. coli under the control of a bicistronic plasmid system in DO-stat fed-batch bioreactor cultivations.

We studied the expression of Bacillus amyloliquefaciens transglutaminase cloned in Escherichia coli BL21(DE3)pLysS harboring the plasmid pBAD/3C/bTGase, a bicistronic expression system, in bioreactor cultivation. Batch and fed-batch controlled as DO-stat strategies were employed for the production of the recombinant enzyme. In 30 h-batch cultivations using Terrific broth (TB), 6 g/L of biomass and 3.12 U/mgprotein of transglutaminase activity were obtained. DO-stat fed-batch cultivations under the control of oxygen concentration (DO-stat) using TB as medium but fed with glucose allowed the increment in biomass formation (17.5 g/L) and enzyme activity (6.43 U/mgprotein). DO-stat fed-batch using mineral medium (M9) and fed with glucose under the same conditions produced even higher enzymatic activity (9.14 U/mgprotein). The pH effect was investigated, and the best enzymatic activity could be observed at pH 8. In all cultivations, the bicistronic system remained stable, with 100% of plasmid-bearing cells. These results show that E. coli bearing bicistronic plasmid constructs to express recombinant TGase could be cultivated in bioreactors under DO-stat fed-batch using mineral medium and it is a promising strategy in future optimizations to produce this important enzyme.

Viability of microencapsulated Lactobacillus acidophilus by complex coacervation associated with enzymatic crosslinking under application in different fruit juices.

The objective of this study was to produce microcapsules containing Lactobacillus acidophilus LA-02 by complex coacervation followed by crosslinking with transglutaminase and to evaluate the effect of their addition on different fruit juices, as well as the probiotic viability of L. acidophilus and its effect on fruit juices during storage. To this end, L. acidophilus was microencapsulated by complex coacervation, followed by crosslinking with transglutaminase at different concentrations. Probiotics, in their free and microencapsulated forms, were added to orange juice and apple juice at concentrations of 10% and 30%. The obtained microcapsules were characterized in terms of morphology. The viability of probiotics and the effects of their addition on fruit juices were assessed and the juices characterized (with respect to pH and total soluble solids) during 63 days of storage at 4 °C. Orange juice proved to be more suitable for the addition of probiotics, and the survival of probiotics was directly related to pH. The microcapsules had a protective effect on L. acidophilus, prolonging their survival, and the crosslinking process proved to be adequate and promising, ensuring probiotic viability. Thus, the complex coacervation process associated with induced enzymatic crosslinking provided protection for L. acidophilus in different fruit juices, showing an adequate methodology for adding probiotics to this adverse food matrix, guaranteeing the survival of L. acidophilus for up to 63 days, and generating products with innovative and promising probiotic appeal.

Evaluation of nuclear NF-κB, transglutaminase2, and ERCC1 as predictors of platinum resistance in testicular tumors

ABSTRACT Purpose: Testicular germ cells tumor (TGCT) are associated with a high cure rate and are treated with platinum-based chemotherapy. However, a group of testicular cancer patients may have a very unfavorable evolution and insensitivity to the main therapeutic agent chemotherapy (CT) cisplatin. The aim of this study was to evaluate the risk of recurrence and overall survival related to the expression of nuclear factor kappa-B (NF-κB), transglutaminase 2 (TG2) and excision repair cross-complementation group 1 (ERCC1) in patients with TGCT treated with platinum combinations. Patients and Methods: A retrospective study was performed with TGCT patients treated with platinum-based chemotherapy. Immunohistochemical analysis was performed and the expression was correlated with clinical and laboratory data. Results: Fifty patients were included, the mean age was 28.4 years (18 to 45), and 76% were non-seminoma. All patients were treated with standard cisplatin, etoposide and bleomycin or cisplatin, and etoposide. Patient's analyzed immunodetection for NF-κB, TG2, and ERCC1 were positive in 76%, 54% and 42%, respectively. Multivariate analysis identified that positive expressions to ERCC1 and NF-κB are independent risk factors for higher recurrence TGCT after chemotherapy (RR 2.96 and 3.16, respectively). Patients with positive expression of ERCC1 presented a poor overall survival rate for 10-year follow (p=0.001). Conclusions: The expression of ERCC1 and NF-κB give a worse prognosis for relapse, and only ERCC1 had an influence on the overall survival of TGCT patients treated with platinum-based chemotherapy. These may represent markers that predict poor clinical outcome and response to cisplatin.