Fabio Salamanca

Characterization of mtDNA Haplogroups in 14 Mexican Indigenous Populations

ABSTRACT In this descriptive study we investigated the genetic structure of 513 Mexican indigenous subjects grouped in 14 populations (Mixteca-Alta, Mixteca-Baja, Otomi, Purépecha, Tzeltal, Tarahumara, Huichol, Nahua-Atocpan, Nahua-Xochimilco, Nahua-Zitlala, Nahua-Chilacachapa, Nahua-Ixhuatlancillo, Nahua-Necoxtla, and Nahua-Coyolillo) based on mtDNA haplogroups. These communities are geographically and culturally isolated; parents and grandparents were born in the community. Our data show that 98.6% of the mtDNA was distributed in haplogroups A1, A2, B1, B2, C1, C2, D1, and D2. Haplotype X6 was present in the Tarahumara (1/53) and Huichol (3/15), and haplotype L was present in the Nahua-Coyolillo (3/38). The first two principal components accounted for 95.9% of the total variation in the sample. The mtDNA haplogroup frequencies in the Purépecha and Zitlala were intermediate to cluster 1 (Otomi, Nahua-Ixhuatlancillo, Nahua-Xochimilco, Mixteca-Baja, and Tzeltal) and cluster 2 (Nahua-Necoxtla, Nahua-Atocpan, and Nahua-Chilacachapa). The Huichol, Tarahumara, Mixteca-Alta, and Nahua-Coyolillo were separated from the rest of the populations. According to these findings, the distribution of mtDNA haplogroups found in Mexican indigenous groups is similar to other Amerindian haplogroups, except for the African haplogroup found in one population.

A molecular analysis by gene expression profiling reveals Bik/NBK overexpression in sporadic breast tumor samples of Mexican females

BackgroundBreast cancer is one of the most frequent causes of death in Mexican women over 35 years of age. At molecular level, changes in many genetic networks have been reported as associated with this neoplasia. To analyze these changes, we determined gene expression profiles of tumors from Mexican women with breast cancer at different stages and compared these with those of normal breast tissue samples.Methods32P-radiolabeled cDNA was synthesized by reverse transcription of mRNA from fresh sporadic breast tumor biopsies, as well as normal breast tissue. cDNA probes were hybridized to microarrays and expression levels registered using a phosphorimager. Expression levels of some genes were validated by real time RT-PCR and immunohistochemical assays.ResultsWe identified two subgroups of tumors according to their expression profiles, probably related with cancer progression. Ten genes, unexpressed in normal tissue, were turned on in some tumors. We found consistent high expression of Bik gene in 14/15 tumors with predominant cytoplasmic distribution.ConclusionRecently, the product of the Bik gene has been associated with tumoral reversion in different neoplasic cell lines, and was proposed as therapy to induce apoptosis in cancers, including breast tumors. Even though a relationship among genes, for example those from a particular pathway, can be observed through microarrays, this relationship might not be sufficient to assign a definitive role to Bik in development and progression of the neoplasia. The findings herein reported deserve further investigation.

Promoting networks between evidence-based medicine and values-based medicine in continuing medical education

BackgroundIn recent years, medical practice has followed two different paradigms: evidence-based medicine (EBM) and values-based medicine (VBM). There is an urgent need to promote medical education that strengthens the relationship between these two paradigms. This work is designed to establish the foundations for a continuing medical education (CME) program aimed at encouraging the dialogue between EBM and VBM by determining the values relevant to everyday medical activities.MethodsA quasi-experimental, observational, comparative, prospective and qualitative study was conducted by analyzing through a concurrent triangulation strategy the correlation between healthcare personnel-patient relationship, healthcare personnels life history, and ethical judgments regarding dilemmas that arise in daily clinical practice.In 2009, healthcare personnel working in Mexico were invited to participate in a free, online clinical ethics course. Each participant responded to a set of online survey instruments before and after the CME program. Face-to-face semi-structured interviews were conducted with healthcare personnel, focusing on their views and representations of clinical practice.ResultsThe healthcare personnels core values were honesty and respect. There were significant differences in the clinical practice axiology before and after the course (P <0.001); notably, autonomy climbed from the 10th (order mean (OM) = 8.00) to the 3rd position (OM = 5.86). In ethical discernment, the CME program had an impact on autonomy (P ≤0.0001). Utilitarian autonomy was reinforced in the participants (P ≤0.0001). Regarding work values, significant differences due to the CME intervention were found in openness to change (OC) (P <0.000), self-transcendence (ST) (P <0.001), and self-enhancement (SE) (P <0.019). Predominant values in life history, ethical discernment and healthcare personnel-patient relation were beneficence, respect and compassion, respectively.ConclusionsThe healthcare personnel participating in a CME intervention in clinical ethics improved high-order values: Openness to change (OC) and Self Transcendence (ST), which are essential to fulfilling the healing ends of medicine. The CME intervention strengthened the role of educators and advisors with respect to healthcare personnel. The ethical values developed by healthcare professionals arise from their life history and their professional formation.

Endothelial nitric oxide synthase gene polymorphism in the Indian and Mestizo populations of Mexico

Endothelium-derived nitric oxide (NO) is an important factor in vasodilation synthesized by endothelial nitric oxide synthase (eNOS). A polymorphism (894 G to T) in exon 7 of the eNOS gene causes the conversion of Glu to Asp in position 298. The Glu298Asp polymorphism has been extensively associated with cardiovascular disease. We determined the Glu298Asp polymorphism frequency in healthy Mexican Mestizo, Huastec, Mayo, and Mayan populations by the endonuclease restriction method. The four populations analyzed were in Hardy-Weinberg equilibrium. Allele frequencies were similar among Mexican populations but different when compared with Caucasians. However, when compared with allele frequencies in Asian populations, Mestizo and Huastec allele frequencies were significantly different. Genotypically, only the Mestizos presented Asp298 homozygosity. The absence of double mutants in Indian populations resembles that in Asians. With these data, we conclude that the low frequency of the eNOS Glu298Asp polymorphism in Indian and Mestizo populations of Mexico is related to the Asian origin of Amerindian groups.

Expression analysis of the SG-SSPN complex in smooth muscle and endothelial cells of human umbilical cord vessels.

Recently, participation of the sarcoglycan (SG)-sarcospan (SSPN) complex in the development of cardiomyopathy in patients with limb-girdle muscular dystrophy has been shown, and presence of the complex in smooth muscle may be important for the contraction/dilation process of vessels. However, there are few studies determining the SG-SSPN complex in vascular smooth muscle and endothelial cells of vessels. In this study, we analyzed by reverse transcriptase-polymerase chain reaction and immunofluorescence the expression of different components of the complex in vein/artery smooth muscle and endothelial cells of the human umbilical cord. By RNA analysis, we observed expression of α-, β-, γ-, δ-, Ε-SG, and SSPN in smooth muscle cells. In endothelial cells, RNA expression was restricted to β-, δ-, Ε-SG, and SSPN. At protein level, we observed in smooth muscle the presence of β-, δ-, Ε-SG, and SSPN. In endothelial cells, immunostaining only evidenced the presence of Ε-SG and SSPN. However, colocalization of SGs and SSPN with dystrophin and utrophin was noted. These results, interestingly, suggest that the SG-SSPN complex may either form with dystrophin or utrophin in smooth muscle cells, and with utrophin in endothelial cells. Additionally, we also observed in some smooth muscle regions the colocalization of the SG-SSPN complex with caveolin, with colocalization being more pronounced between Ε-SG-SSPN and caveolin in endothelial cells.

Frequency of haplotypes in the beta globin gene cluster in a selected sample of the mexican population

Five polymorphic restriction enzyme sites in the beta globin gene cluster (HindIII Gγ‐Hind III Aγ‐, Ava IIINV−2β‐and Hpa I and Bam HI 3′β‐globin gene) were studied in individuals from 13 families: 13 homozygote patients for sickle cell anemia, two double heterozygotes (one SC and one S/βThal), 35 AS heterozygotes (23 parents and 12 siblings), one father (A/βThal), and three normal siblings. In addition, 17 normal unrelated Mexican subjects were studied. All subjects were from the state of Veracruz on the coast of the Gulf of Mexico. The Southern blot technique was used. Fifteen haplotypes were identified in the 142 chromosomes. Five were the most frequent: two haplotypes, (+−+++) (52.4%) and (−−+−+) (19.0%) were associated with βS chromosomes; two haplotypes, (−−+++) (38.2%) and (−−−++) (19.7%), were linked with βA chromosomes, and the fifth (−−++−) was present in both types of chromosomes. Haplotype (+−+++) corresponded to the Bantu or Senegal type. With Hinc II analysis after PCR amplification in both the 5′ and 3′ regions of the ψβ‐globin gene, it was possible to distinguish between these African types, as in the former both restriction sites are absent. This analysis was done in 23 βS and 10 βA subjects. All βS chromosomes disclosed the Bantu type, while βA were similar to Caucasians. Bantu and Benin haplotypes have been found with high frequency in African populations, indicating the great influence of African genes in the population of the Mexican coasts. In addition, two previously unidentified haplotypes were found: (++−−+) and (−++++). These can be explainded by crossing‐over events and/or by new mutations.

Expression and function of utrophin associated protein complex in stretched endothelial cells: dissociation and activation of eNOS.

Several studies have emphasized the relevance of dystrophin-associated protein complex (DAPC) to maintain the vascular function. Previously we postulated the presence of an utrophin associated protein complex (UAPC) in endothelium from umbilical cord vessels. In the present work, we demonstrate that utrophin (UTR) indeed forms a complex, with beta-dystroglycan (DG), epsilon-sarcoglycan (SG), caveolin-1 (cav-1), and endothelial nitric oxide synthase (eNOS) in human umbilical vein endothelial cells (HUVEC) by co-immunoprecipitation analysis. Additionally, we observed an increment in the protein levels of epsilon-SG, beta-DG, UTR and cav-1 after mechanical stretching. Interestingly, this stimulus also induced eNOS up-regulation, activation and release from the UAPC, and led to a significant increase in nitric oxide (NO) production. Finally, we propose that UAPC in HUVECs may play an important role in the regulation of vascular tone.

Identification of novel mutations in the RB1 gene in Mexican patients with retinoblastoma

Retinoblastoma (RB) is a childhood tumor of the eye with an average incidence of one case in every 15,000-20,000 live births and occurs in sporadic or hereditary form. This cancer results from loss or inactivation of the RB1 gene located at 13q14.1. This gene encodes for a 110 Kd nuclear phosphoprotein (pRB) that plays a major role in cell proliferation control. Different types of mutations in the RB1 gene have been reported, but point mutations are the most common. There are no molecular studies on RB1 gene mutation in Mexican patients. In this study, 19 patients with bilateral or unilateral RB were analyzed. Genetic and cytogenetic studies were carried out. Detection of RB1 gene mutations was done using single-strand conformational polymorphism (SSCP). Five conformational polymorphisms were identified in different exons. In all cases, SSCP sequence showed new non-described mutations that produced a frameshift on the open reading frame. The identification of mutations in the RB1 gene contributes to basic knowledge of this neoplasia and permits the possibility to offer adequate genetic counseling to relatives at risk.

Using linear-bilinear models for studying gene expression × treatment interaction in microarray experiments

In microarray experiments, the global and the specific gene expression in the two-way table of gene x treatments (or tissues) can be studied using linear-bilinear models that incorporate the main effects of genes (G), treatment (T), and gene x treatment interaction (G x T). The plot of the first two axes obtained from the singular value decomposition of the bilinear (multiplicative) term of these models (biplot) facilitates the interpretation of the gene expression patterns. In this study, two microarray datasets were used to illustrate how two linear-bilinear models, the additive main effect and multiplicative interaction (AMMI) and the treatment regression model (TREG) and their biplots can be used to determine the overall gene expression pattern across treatments (or tissues) and for specific treatments. Dataset 1 had 5,339 genes and the objective was to identify genes with modified expression during maize (Zea mays) seed development in response to different parental ploidy levels. In Dataset 2, the aim was to study gene expression in 15 tissue samples with different levels of development of breast cancer when compared with the expression of the genes in noninfected tissues. The results from the analyses of Dataset 1 showed that the biplots of the AMMI and TREG models allow identification of subsets of genes and treatments with noncrossover G x T interaction or with important levels of crossover G x T. Results from Dataset 2 showed that the TREG model and its biplot facilitates the identification of genes with high expression in all tumor cells. Also, the TREG biplots allowed identification of subsets of genes with a low level of gene x tissue crossover interaction.

PCR–PRINS–FISH analysis of structurally abnormal sex chromosomes in eight patients with Turner phenotype

According to cytogenetic analysis, about 50% of Turner individuals are 45,X. The remaining cases have a structurally abnormal X chromosome or are mosaics with a second cell line containing a normal or abnormal sex chromosome. In these mosaics, approximately 20% have a sex marker chromosome whose identity cannot usually be determined by classical cytogenetic methods, requiring the use of molecular techniques. Polymerase chain reaction (PCR), primed in situ labeling (PRINS), and fluorescence in situ hybridization (FISH) analyses were performed in 8 patients with Turner syndrome and 45,X mosaic karyotypes to determine the origin and structure of the marker chromosome in the second cell line. Our data showed that markers were Y‐derived in 2 patients and X‐derived in the remaining 6 patients. We were also able to determine the breakpoints in the two Y chromosomes. The use of cytogenetic and molecular techniques allowed us to establish unequivocally the origin, X or Y, of the marker chromosomes in the 8 patients with Turner phenotype. This study illustrates the power of resolution and utility of combined cytogenetic and molecular approaches in some clinical cases.