Veratridine-sensitive Na+ channels regulate human sperm fertilization capacity.

AIMS: The sperm plasma membrane contains specific ion channels and transporters that initiate changes in Ca2+, Na+, K+ and H+ ions in the sperm cytoplasm. Ion channels are key regulators of the sperm membrane potential, cytoplasmic Ca2+ and intracellular pH (pHi), which leads to regulate motility, capacitation, acrosome reaction and other physiological processes crucial for successful fertilization. Expression of epithelial sodium channels (ENaC) and voltage-gated sodium channels (Nav) in human spermatozoa has been reported, but the role of Na+ fluxes sodium channels in the regulation of sperm cell function remains poorly understood. In this context, we aimed to analyze the physiological role of Nav channels in human sperm. MAIN METHODS: Motility and hyperactivation analysis was conducted by CASA analysis. Flow cytometry and spectrophotometry approaches were carried out to measure Capacitation, Acrosome reaction, immunohistochemistry for Tyr-residues phosporylation, [Ca2+]i levels and membrane potential. KEY FINDINGS: Functional studies showed that veratridine, a voltage-gated sodium channel activator, increased sperm progressive motility without producing hyperactivation while the Nav antagonist lidocaine did induce hyperactivated motility. Veratridine increased protein tyrosine phosphorylation, an event occurring during capacitation, and its effects were inhibited in the presence of lidocaine and tetrodotoxin. Veratridine had no effect on the acrosome reaction by itself, but was able to block the progesterone-induced acrosome reaction. Moreover, veratridine caused a membrane depolarization and modified the effect of progesterone on [Ca2+]i and sperm membrane potential. SIGNIFICANCE: Our results suggest that veratridine-sensitive Nav channels are involved on human sperm fertility acquisition regulating motility, capacitation and the progesterone-induced acrosome reaction in human sperm.

Decatropis bicolor (Zucc.) Radlk essential oil induces apoptosis of the MDA-MB-231 breast cancer cell line.

BACKGROUND: Decatropis bicolor (Zucc.)Radlk is a plant that has been traditionally used for the treatment of breast cancer in some communities of Mexico. So, the aim of this study was to determine the cytotoxic and apoptotic effect of the essential oil of Decatropis bicolor against breast cancer cell line, MDA-MB-231. METHODS: The essential oil obtained from hydrodestillation of leaves of Decatropis bicolor was studied for its biological activity against breast cancer cells MDA-MB-231 by MTT assay, Hematoxylin-eosin stain, Annexin V-FITC, TUNEL and western blot assays and for its chemical composition by GC-MS. RESULTS: The results showed a relevant cytotoxic effect of the essential oil towards MDA-MB-231 cells in a dose- and time- dependent manner, with an IC50 of 53.81 ± 1.691 µg/ml but not in the epithelial mammary cell line MCF10A (207.51 ± 3.26 µg/ml). Morphological examination displayed apoptotic characteristics in the treated cells like cell size reduction, membrane blebbing and apoptotic bodies. In addition, the apoptotic rate significantly increased as well as DNA fragmentation and western blot analysis revealed that the essential oil induced apoptosis in the MDA-MB-231 cells via intrinsic pathways due to the activation of Bax, caspases 9 and 3. Phytochemical analysis of the Decatropis bicolor essential oil showed the presence of twenty-three compounds. Major components of the oil were 1,5-cyclooctadiene,3-(methyl-2)propenyl (18.38 %), ß-terpineol (8.16 %) and 1-(3-methyl-cyclopent-2-enyl)-cyclohexene (6.12 %). CONCLUSIONS: This study suggests that essential oil of Decatropis bicolor has a potential cytotoxic and antitumoral effect against breast cancer cells, with the presence of potential bioactive compounds. Our results contribute to the validation of the anticancer activity of the plant in Mexican traditional medicine.

rs3918242 MMP9 gene polymorphism is associated with myocardial infarction in Mexican patients.

Several studies have demonstrated that matrix metalloproteinases (MMPs) play a major role in atherosclerotic plaque disruption and lead to myocardial infarction (MI). We investigated the association between the MMP1 -1607 1G/2G (rs1799750), MMP3 -1612 5A/6A (rs3025058), and MMP9 -1562 C/T (rs3918242) polymorphisms and the risk of developing MI in a Mexican mestizo cohort. The genotype analysis was performed using the restriction fragment length polymorphism-polymerase chain reaction technique in a group of 236 patients with a history of MI and 285 healthy controls. Similar distributions of rs1799750 and rs3025058 were observed in both groups; however, the MMP9 rs3918242 T allele and the CT genotype were associated with the risk of developing MI (OR = 2.32, pC = 0.02 and OR = 2.40, pC = 0.02, respectively). Multiple logistic analysis was performed between MI patients and controls to estimate the risk, and after adjusting for identified risk factors, the CT + TT genotypes of MMP9 rs3918242 were found to be significantly associated with increased risk of developing MI than those with the CC genotype (OR = 2.88, P < 0.01). In summary, our results reveal that the rs3918242 polymorphism of the MMP9 gene plays a major role in the risk of developing MI.

Antinociceptive, genotoxic and histopathological study of Heliopsis longipes S.F. Blake in mice.

ETHNOPHARMACOLOGICAL RELEVANCE: H. longipes S.F. Blake (Asteraceae) is a Mexican plant, whose roots are traditionally used as a condiment, as a mouth anesthetic, and as an antiparasitic. Affinin is the alkamide present in higher amounts in the roots of H. longipes. AIM OF THE STUDY: To date, there are no published studies regarding the relation between the analgesic properties, in vivo cytotoxicity, and DNA-damaging potential of H. longipes ethanol extract (HLEE). MATERIALS AND METHODS: The HLEE was chromatographically fingerprinted to validate its affinin contents. Biological evaluation was conducted in sets of 6-8 CD1(+) mice. Antinociceptive effect was evaluated using the writhing and hot-plate tests, and mutagenic and cytotoxic effects were evaluated with micronucleous test in CD1(+) mice. For histopathological studies, biological samples from liver, heart, kidneys, spleen, lung, and brain were collected and stained. RESULTS: Oral administration of HLEE (3-100 mg/kg) produced a dose-dependent antinociceptive effect in both assays. In micronucleus assay, the variability in the number of micronucleated polychromatic erythrocytes (MNPE) induced, and PE/NE index, the ratio of polychromatic erythrocytes with respect to the number of normochromatic erythrocytes induced by HLEE in the evaluated schedule, were small and nonsignificant. After histopathological results, HLEE showed polioencephalomalacia with 1000 mg/kg dose. CONCLUSIONS: This work provides evidence that HLEE exerts analgesic effects, with no genotoxic effects in vivo. These findings would be an important contribution to explain the use of H. longipes root as an effective analgesic in traditional medicine, and to establish for the first time the absence of genotoxic and cytotoxic effects of the root in bioactive doses in vivo.

Association of HLA DRB1 alleles with juvenile idiopathic arthritis in Mexicans.

OBJECTIVE: The aim of the study was to investigate association between HLA class II alleles and juvenile idiopathic arthritis (JIA) in Mexican patients. PATIENTS AND METHODS: We typed 120 patients with JIA and 99 healthy controls for HLA class II alleles were performed by PCR-SSO. Differences between the whole group of JIA and its subtypes and controls were calculated by using the Xi2; p-values were corrected (pc) with Bonferroni's test. RESULTS: The alleles HLA-DRB1*01 (pc= 0.00083) and HLA-DRB1*04 (pc=0.0049) were strongly associated with systemic JIA, while HLA-DRB1*11 and HLA-DRB1*14 were found to have decreased frequencies in the patients with systemic JIA compared to the controls. Two alleles were found to have increased frequencies with JIA oligoarthritis subgroup, HLA-DRB1*11 (p=0.01, pc=NS) and HLA-DRB1*13 (p=0.01, pc=NS). The HLA-DRB1*04 was found increased frequencies with susceptibility for RF negative and RF positive polyarthritis JIA subgroups (p correction resulted in loss of significance). In contrast two alleles HLA-DRB1*07 and HLA-DRB1*14 were found decreased frequencies only patients RF positive polyarthritis JIA subgroup compared to the controls (pc=NS). CONCLUSION: The profile of HLA-DRB1 alleles associations in Mexican with JIA were somewhat distinct from association typically found in Caucasians.

Interaction between Heliopsis longipes extract and diclofenac on the thermal hyperalgesia test.

Heliopsis longipes is an herbaceous plant found in Mexico, used traditionally for its analgesic and anesthetic activities. Plant extracts in combined use with synthetic drugs may represent a therapeutic advantage for the clinical treatment of pain, allowing the use of lower doses, and limiting side-effects. Therefore, the main objective of this study was to determine the possible pharmacological interaction between Heliopsis longipes ethanolic extract (HLEE) and diclofenac in the Hargreaves model of thermal hyperalgesia in the mouse. HLEE, diclofenac or fixed-dose ratio HLEE-diclofenac combinations were administered systemically to mice and the antihyperalgesic effect was evaluated using the thermal hyperalgesia test. All treatments produced a dose-dependent antihyperalgesic effect. ED(30) values were estimated for all the treatments and an isobologram was constructed. The derived theoretical ED(30) value for the HLEE-diclofenac combination was 54.4+/-9.4 mg/kg body wt, significantly higher than the actually observed experimental ED(30) value, 8.6+/-4.0 mg/kg body wt. This result corresponds to synergistic interaction between HLEE and diclofenac in the Hargreaves model of thermal hyperalgesia. Data suggest that low doses of the HLEE-diclofenac combination can interact synergistically at the systemic level and that this association may therefore represent a therapeutic advantage for the clinical treatment of inflammatory pain.

HLA Genes in Mayos Population from Northeast Mexico.

HLA class I and class II alleles have been studied in 60 unrelated people belonging to Mayos ethnic group, which lives in the Mexican Pacific Sinaloa State. Mayos HLA profile was compared to other Amerindians and worldwide populations' profile. A total of 14,896 chromosomes were used for comparisons. Genetic distances between populations, Neigbour-Joining dendrograms and correspondence analyses were performed to determine the genetic relationship among population. The new specific Mayo HLA haplotypes found are: HLA-A*02-B*35-DRB1*1406-DQB1*0301; HLA-A*02-B*48-DRB1*0404-DQB1*0302; HLA-A*24-B*51-DRB1*0407-DQB1*0302 and HLA-A*02-B*08-DRB1*0407-DQB1*0302. However, the typical Meso American HLADRB1*0407 represents a 40% of all DRB1 alleles. While common HLA characteristics are found in Amerindian distant ethnic groups, still new group specific HLA haplotypes are being found, suggesting that a common founder effect (i.e. high DRB1*0407) is noticed. Moreover, new HLA haplotypes are almost certainly appearing along time probably due to specific pathogen (?) selection for diversity. Mayo language is close to the Tarahumara one (another geographically close group); notwithstanding both groups are not genetically close according to our results, showing again the different evolution of genes and languages, which do not correlate. Finally, Sinaloa is one of the Mexican States in which more European genes are found. However, the results presented in this paper, where no European HLA genes are seen in Mayos, should have a bearing in establishing transplant programs and in HLA and disease studies.

Hydrogen-bonding patterns of two dihydroxylactone derivatives.

In the hydrogen-bonding networks of 8-hydroxy-5-hydroxymethyl-3,6-dioxatricyclo[6.3.1.0(1.5)]dodecan-2-one and 5,7-bis(hydroxymethyl)-3,6-dioxatricyclo[5.3.1.0(1.5)]undecan-2-one, both C11H16O5, layers and double strands, respectively, lead to the formation of chains connected by hydroxy-to-hydroxy contacts, where the hydroxymethyl group, present in both structures, acts as a donor. The secondary structures differ in the hydrogen bonding of these chains via the second hydroxy group, which is involved in hydroxy-to-carbonyl and hydroxy-to-hydroxy bonds, respectively.

HLA-DRB1 alleles encoding the "shared epitope" are associated with susceptibility to developing rheumatoid arthritis whereas HLA-DRB1 alleles encoding an aspartic acid at position 70 of the beta-chain are protective in Mexican Mestizos.

The risk to develop rheumatoid arthritis (RA) has been associated with the presence of HLA-DRB1 alleles encoding the "shared epitope" (SE). Additionally, HLA-DRB1 alleles encoding an aspartic acid at position 70 (D70+ ) have been associated with protection against the development of RA. In this study we tested the association between either SE or D70+ and rheumatoid arthritis in Mexican Mestizos. We included 84 unrelated Mexican Mestizos patients with RA and 99 unrelated healthy controls. The HLA-typing was performed by PCR-SSO and PCR-SSP. We used the chi-squared test to detect differences in proportions of individuals carrying at least one SE or D70+ between patients and controls. We found that the proportion of individuals carrying at least one HLA-DRB1 allele encoding the SE was significantly increased in RA cases as compared to controls (p(c) = 0.0004, OR = 4.1, 95% CI = 2.2-7.7). The most frequently occurring allele was HLA-DRB1*0404 (0.161 vs 0.045). Moreover, we observed a significantly increased proportion of HLA-DRB1 SE+ cases with RF titers above the median (p = 0.005). Conversely, the proportion of individuals carrying at least one HLA-DRB1 allele encoding the D70+ was significantly decreased (p(c) = 0.004, OR = 0.4, 95% CI 0.2-0.7) among RA patients compared with controls. In conclusion, the SE is associated with RA in Mexican Mestizos as well as with the highest titers of RF.

Biological relevance of the polymorphism in the CCR5 gene in refractory and non-refractory rheumatoid arthritis in Mexicans.

OBJECTIVE: The aim of this study was to analyze the frequencies of the CCR5 delta 32 deletion and HLA class II alleles in Mexican Amerindian populations and its relevance in the development and severity of RA. METHODS: We studied 212 Mexican Mestizo subjects (40 patients with refractory RA, 102 patients with non-refractory RA and 70 healthy individuals). At the same time, to evaluate the ethnicity of the CCR5 delta 32 deletion we also studied 192 individuals from three Mexican Amerindian populations (70 Mayo (Capomo) individuals, 61 Teenek individuals, and 61 Mazatecan Indians). The delta 32 deletion in the CCR5 structural gene and HLA-DRB1 were determined by a PCR-SSP and a PCR-SSO procedure, respectively. RESULTS: In the non-refractory RA group the CCR5 delta 32 gene frequency was 0.019 and the following genotype frequencies were observed: CCR5/CCR5 = 98.0%, CCR5/CCR5 delta 32 = 1.9% and CCR5 delta 32/CCR5 delta = 1.0%. In the refractory RA group the CCR5 delta 32 gene frequency was 0.025 and the genotype distribution was similar to that in the non-refractory RA group. The deletion was not detected in the Mexican Mestizo healthy group, or among the Teenek and Mayo Amerindians, all being individuals homozygous for the wild type allele. In the Mazatecan group the deletion frequency was 1.6% (g.f. = 0.016). We observed a significant increase in the frequency of the DRB1*07 allele in severe RA patients in relation to the non-severe RA group (p = 0.02, OR = 5.65, 95% CI = 0.95-43.05). CONCLUSION: Our results suggest that the CCR5 delta 32 deletion is not common in Mexican Amerindian populations and this study does not support an important role of CCR5 delta 32 in the pathogenesis of RA or a severe form of the disease in Mexicans.