Limonin mitigates cardiometabolic complications in rats with metabolic syndrome through regulation of the IRS-1/GLUT4 signalling pathway.

Limonin is a natural triterpenoid isolated from citrus fruit. In the present study, we examined the effects of limonin on cardiometabolic alterations in diet-induced metabolic syndrome. Metabolic syndrome was induced in rats by feeding them a high-fat (HF) diet plus 15% fructose in drinking water for 16 weeks. They were treated with limonin (50 or 100 mg/kg) (n = 8/group) for the final 4 weeks. Increases in body weight (BW), fasting blood glucose (FBG), serum insulin, total cholesterol (TC), blood pressure (BP), liver fat accumulation, and adipocyte hypertrophy, as well as oral glucose tolerance in rats with metabolic syndrome were alleviated by limonin treatment (p < 0.05). Limonin improved ejection fraction and left ventricular (LV) hypertrophy, and reduced angiotensin converting enzyme (ACE) activity and angiotensin II (Ang II) concentration in rats with metabolic syndrome (p < 0.05). It also reduced plasma tumour necrosis factor (TNF)-α, interleukin (IL)- 6, leptin, malonaldehyde (MDA), and superoxide generation, and increased catalase activity in rats with metabolic syndrome compared to controls (p < 0.05). Downregulation of insulin receptor substrate 1 (IRS-1) and glucose transporter type 4 (GLUT4) protein expression in epididymal fat pads and cardiac, liver, and gastrocnemius tissues was present in metabolic syndrome, and these were restored by limonin treatment (p < 0.05). In conclusion, limonin shows a potential effect in alleviating symptoms and improving cardiometabolic disorders. These beneficial effects are linked to the reduction of the renin-angiotensin system, inflammation, oxidative stress, and improvement of IRS-1/GLUT4 protein expression in the target tissue.

Vasoactive Properties of a Cocoa Shell Extract: Mechanism of Action and Effect on Endothelial Dysfunction in Aged Rats.

Cocoa has cardiovascular beneficial effects related to its content of antioxidant phytochemicals. Cocoa manufacturing produces large amounts of waste, but some by-products may be used as ingredients with health-promoting potential. We aimed to investigate the vasoactive actions of an extract from cocoa shell (CSE), a by-product containing theobromine (TH), caffeine (CAF) and protocatechuic acid (PCA) as major phytochemicals. In carotid and iliac arteries from 5-month and 15-month-old rats, we investigated CSE vasoactive properties, mechanism of action, and the capacity of CSE, TH, CAF and PCA to improve age-induced endothelial dysfunction. Vascular function was evaluated using isometric tension recording and superoxide anion production by dihydroethidium (DHE) staining and confocal microscopy. CSE caused endothelium-dependent vasorelaxation, blocked by L-NAME, but not indomethacin, regardless of sex, age, or vessel type. CSE maximal responses and EC50 were significantly lower compared to acetylcholine (ACh). Arterial preincubation with CSE, TH, CAF or PCA, significantly reduced the number of vascular DHE-positive cells. Compared to adult males, iliac arteries from aged males exhibited reduced ACh concentration-dependent vasodilatation but larger CSE responses. In iliac arteries from aged male and female rats, preincubation with 10-4 M CSE and PCA, but not TH or CAF, improved ACh-relaxations. In conclusion, CSE has vasodilatory properties associated with increased nitric oxide bioavailability, related to its antioxidant phytochemicals, being particularly relevant PCA. Therefore, CSE is a potential food ingredient for diseases related to endothelial dysfunction.

Combination of Melatonin and Small Molecules Improved Reprogramming Neural Cell Fates via Autophagy Activation.

Reprogramming cell fates towards mature cell types are a promising cell supply for treating degenerative diseases. Recently, transcription factors and some small molecules have turned into impressive modulating elements for reprogramming cell fates. Melatonin, a pineal hormone, has neuroprotective functions including neural stem cell (NSC) proliferative and differentiative modulation in both embryonic and adult brain. We developed a protocol that could be implemented in the direct reprogramming of human skin fibroblast towards neural cells by using histone deacetylase (HDAC) inhibitor, glycogen synthase kinase-3 (GSK3) inhibitor (CHIR99021), c-Jun N-terminal kinase (JNK) inhibitor, rho-associated protein kinase inhibitor (Y-27632), cAMP activator, and melatonin treatment. We found that melatonin enhanced neural-transcription factor genes expressions, including brain-specific homeobox/POU domain protein 2 (BRN2), Achaete-Scute Family BHLH transcription Factor 1 (ASCL1), and Myelin Transcription Factor 1 Like (MYT1L). Melatonin also increased the expression of different neural-specific proteins such as doublecortin (DCX), Sex determining region Y-box 2 (Sox2), and neuronal nuclei (NeuN) compared with other five small molecules (valproic acid (VPA), CHIR99021, Forskolin, 1,9 pyrazoloanthrone (SP600125), and Y-27632) combination in the presence and absence of melatonin. A noticeable upregulation of autophagy proteins (microtubule-associated protein 1A/1B-light chain 3 (LC3) and Beclin-1) were seen in the melatonin treatment during the induction period while these were reverted in the presence of L-leucine, an autophagy inhibitor. In addition, the expression of NeuN was also significantly reduced by L-leucine. Collectively, our findings revealed an activation of autophagy during neural induction; melatonin enhanced reprogramming efficiency for neuron induction through the modulation of autophagy activation.

Sex Differences in Placental Protein Expression and Efficiency in a Rat Model of Fetal Programming Induced by Maternal Undernutrition.

Fetal undernutrition programs cardiometabolic diseases, with higher susceptibility in males. The mechanisms implicated are not fully understood and may be related to sex differences in placental adaptation. To evaluate this hypothesis, we investigated placental oxidative balance, vascularization, glucocorticoid barrier, and fetal growth in rats exposed to 50% global nutrient restriction from gestation day 11 (MUN, n = 8) and controls (n = 8). At gestation day 20 (G20), we analyzed maternal, placental, and fetal weights; oxidative damage, antioxidants, corticosterone, and PlGF (placental growth factor, spectrophotometry); and VEGF (vascular endothelial growth factor), 11ß-HSD2, p22phox, XO, SOD1, SOD2, SOD3, catalase, and UCP2 expression (Western blot). Compared with controls, MUN dams exhibited lower weight and plasma proteins and higher corticosterone and catalase without oxidative damage. Control male fetuses were larger than female fetuses. MUN males had higher plasma corticosterone and were smaller than control males, but had similar weight than MUN females. MUN male placenta showed higher XO and lower 11ß-HSD2, VEGF, SOD2, catalase, UCP2, and feto-placental ratio than controls. MUN females had similar feto-placental ratio and plasma corticosterone than controls. Female placenta expressed lower XO, 11ß-HSD2, and SOD3; similar VEGF, SOD1, SOD2, and UCP2; and higher catalase than controls, being 11ß-HSD2 and VEGF higher compared to MUN males. Male placenta has worse adaptation to undernutrition with lower efficiency, associated with oxidative disbalance and reduced vascularization and glucocorticoid barrier. Glucocorticoids and low nutrients may both contribute to programming in MUN males.

Genetic Polymorphisms of Vitamin D Receptor Gene are Associated with Cervical Cancer Risk in Northeastern Thailand.

OBJECTIVE: This study aimed to explore whether VDR polymorphisms (Fok1, Apa1 and Taq1) are associated to the cervical cancer in Thai population. MATERIALS AND METHODS: Subjects of 204 cervical cancer patient and 204 age-matched healthy control were enrolled in the case-control study. VDR polymorphisms were detected by using real-time PCR. Haplotype analysis of three loci was applied to the obtained genotypes. RESULTS: Significantly increased risk for cervical cancer was observed in carriers of TT genotype (p = 0.0388) and T allele (p = 0.0357) of Fok1 and TC genotype (p = 0.0001), CC genotype (p = 0.0160) and the C allele of Taq1 (p = 0.0001). Haplotype analyses revealed a significant correlation between C-T-C, T-G-C and T-T-C haplotypes and elevated risk for cervical cancer (OR = 2.06; 95%CI = 1.06-4.00; p = 0.0313, OR = 2.15; 95%CI = 1.22-3.80; p = 0.0078 and OR = 2.81; 95%CI = 1.53-5.16; p = 0.0006, respectively). Furthermore, haplotype carrying C allele of Taq1 (C-G-C + C-T-C + T-G-C + T-T-C) significantly increased cervical cancer risk with OR of 1.92 (95%CI = 1.32-2.79, p = 0.0006). CONCLUSION: Our finding revealed an association between VDR polymorphisms and cervical cancer risk. Taq1 C allele might be a molecular marker for cervical cancer development.
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First trimester elevations of hematocrit, lipid peroxidation and nitrates in women with twin pregnancies who develop preeclampsia.

Twin pregnancies are considered a risk factor for preeclampsia, an obstetric complication with high maternal and infant morbi-mortality. We hypothesize that alterations in maternal hematocrit, plasma lipid peroxidation and nitrates in the first trimester of pregnancy are associated with preeclampsia development in twin pregnancies. Blood samples were extracted from 102 healthy women with twin pregnancies at tenth week of gestation to assess hematological parameters and plasma levels of malondialdehyde and nitrates. Logistic regression model showed an association between red blood cells (OR = 38.8; p-value = 0.009), hematocrit (OR = 1.6; p-value = 0.017), malondialdehyde (OR = 1.5; p-value = 0.002), and nitrates (OR = 1.1; p-value = 0.045) and preeclampsia development. These parameters are potential biomarkers for early preeclampsia detection in twin pregnancies. Future research is needed to assess their value in predictive algorithms.

Development and Validation of a Questionnaire to Assess Adherence to the Healthy Food Pyramid in Spanish Adults.

We aimed to design and validate a new questionnaire of adherence to healthy food pyramid (HFP) (AP-Q), to improve previous instruments. The questionnaire was self-administered and included 28 questions from 10 categories (physical activity, health habits, hydration, grains, fruits, vegetables, oil type, dairy products, animal proteins, and snacks). A population of 130 Spanish adults answered it, obtaining scores from each category and a global score of HFP adherence (AP-Q score). Validation was performed through principal components analysis (PCA) and internal consistency by Cronbach's alpha. AP-Q was also externally validated with Kidmed-test, answered by 45 individuals from the cohort. The global AP-Q score was 5.1 ± 1.3, with an internal consistency of 64%. The PCA analysis extracted seven principal components, which explained 68.5% of the variance. The global AP-Q score was positively associated with Kidmed-test score. Our data suggest that AP-Q is a complete and robust questionnaire to assess HFP adherence, with several advantages: easy to complete, cost-effective, timesaving and has the competency to assess, besides diet, several features affecting health status, lacking in other instruments. We suggest that AP-Q could be useful in epidemiological research, although it requires additional calibration to analyze its reproducibility and validation in other populations.

Genetic Polymorphisms of the Human Cytochrome P450 1A1 (CYP1A1) and Cervical Cancer Susceptibility among Northeast Thai Women.

BACKGROUND: CYP1A1 is an enzyme in phase I of the cytochrome P450 (CYP) superfamily, and plays a key role in detoxification of carcinogens. Host genetic predisposition in the CYP1A1 may be associated with an increased susceptibility to cervical cancer.The study aimed to evaluate four common polymorphisms of the CYP1A1 and cervical cancer susceptibility among Northeast Thai women. METHODS: A case-control study was conducted involving 204 patients with squamous cell cervical cancer (SCCA) and 204 age-matched healthy controls. DNA was extracted from peripheral blood leucocytes. CYP1A1 m1, m3, and m4 genotypes were detected using PCR-RFLP, whereas the CYP1A1 m2 genotype was investigated using real-time PCR. Haplotype analysis was performed using PHASE algorithm version 2.1.1. RESULTS: CYP1A1 m3 was monomorphic. Association between the common CYP1A1 polymorphisms, m1 and m2, and cervical cancer risk was not observed (p>0.05), nor was any association found between the m1-m2-m4 haplotype and cervical cancer risk (p>0.05). Interestingly, the CA genotype of CYP1A1 m4 was observed in 30.88% of the cervical cancer patients but was absent in healthy controls. CONCLUSION: Our results demonstrated a possible involvement of the CYP1A1 m4 polymorphism but no other common polymorphisms (viz., m1, m2, and m3) in the risk for cervical cancer.This finding may be useful when screening for risk of cervical cancer among Northeast Thai women.
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Genetic Polymorphism of the Glutathione S-transferase Pi 1 (GSTP1) and Susceptibility to Cervical Cancer in Human Papilloma Virus Infected Northeastern Thai Women

Objective: We aimed to investigate any association between a genetic polymorphism of the detoxification GSTP1 gene and risk of cervical cancer in northeastern Thailand. Materials and Methods: Genotyping of GSTP1 was performed for 198 squamous cell cervical cancer (SCCA) patients and 198 age-matched healthy controls with the PCR-RFLP method. Results: The respective frequencies of the G allele were 0.33 and 0.26 in the controls and cases, the difference being significant (OR = 0.69 [95% CI: 0.50-0.95, p=0.0192]). Among women infected with high-risk types of HPV, being a heterozygous carrier was associated with a reduced risk of cervical cancer (adjusted OR = 0.32 [95% CI: 0.12-0.91, p=0.031]). Similarly, a decreased risk was observed in heterozygous women with a non-smoking partner (adjusted OR = 0.27 [95% CI: 0.09-0.83, p=0.023]). Conclusions: GSTP1 polymorphism could influence susceptibility to cervical cancer among northeast Thai women; either as a independent factor or in combination with high-risk HPV infection. Dual-testing of HPV and the GSTP1 might prove an effective screening tool for cervical cancer.

Haplotype Analysis of MDR1 and Risk for Cervical Cancer in Northeastern Thailand

Objective: The aim of this study was to investigate the association between genotype and haplotype of MDR1 (C1236T, G2677T/A and C3435T) and the risk for cervical cancer in Northeastern Thai women. Methods: An age-matched case-control study involving squamous cell cervical cancer (SCCA) patients (n=204) and healthy controls (n=204) was enrolled for MDR1 genotyping by real-time PCR method. Results: The genotype distribution of MDR1 in both patients and controls was not significantly different (p>0.05). The haplotype analysis showed that T-T-T was the most common haplotype in this population. Significantly increased risk of cervical cancer was observed in carriers of T-T-C and C-G-T haplotypes with ORs of 1.86 (95%CI=1.02-3.39, p=0.0416) and 2.00 (95%CI=1.18-3.40, p=0.0140), respectively. Analysis of 2677-3435 haplotype showed increased risk for cervical cancer in G-T (OR=1.55; 95% CI=1.12-2.13, p=0.0432) and T-C (OR=1.91; 95%CI=1.05-3.47, p=0.0325). Conclusion: The results provide evidence that haplotype of MDR1 may be an important risk factor for cervical cancer development in Northeastern Thai women.

Preliminary Study of the GSTM1 Null Polymorphism and History of Tobacco Smoking among Oral Cancer Patients in Northeastern Thailand.

Risks with GSTM1 genotypes and potential roles of smoking in the susceptibility to oral squamous cell carcinoma (OSCC) were studied in Northeastern Thailand. Study subjects were 79 histologically-confirmed OSCC cases (31 men, 48 women) and 79 age- and sex-matched healthy controls ranging in age from 25 to 84 years. GSTM1 genotyping was achieved by two independent PCR assays. The GSTM1 null allele and the homozygous genotype did not increase risk of OSCC vs the wild type allele and the remaining genotypes. When the focus was on the smoking habit, male subjects who smoked ≥10 or ≥35 years were at significantly increased risk for OSCC with adjusted ORs of 4.88 [95%CI, 1.41-16.87, p=0.012] or 4.94 [95%CI, 1.62-15.12, p=0.005], respectively. A higher risk for OSCC was found for smoking amount; those who smoked >5 or >10 pack-years were at a higher risk with adjusted OR of 4.46 [95%CI; 1.45-13.74, p=0.009] or 3.89 [95%CI; 1.34-11.28, p=0.012], respectively. There are certain smoking patterns that give greater risks and thus both smoking duration and pack-years should be taken into consideration in tobacco related cancer prevention.