Oxidative stress and mitochondrial dysfunction have been implicated in the pathology of HD, however the precise mechanisms by which mutant huntingtin modulates levels of oxidative damage in turn resulting in mitochondrial dysfunction are not known. We hypothesize that mutant huntingtin increases oxidative mtDNA damage leading to mitochondrial dysfunction. We measured nuclear and mitochondrial DNA lesions and mitochondrial bioenergetics in the STHdhQ7 and STHdhQ111 in vitro striatal model of HD. Striatal cells expressing mutant huntingtin show higher basal levels of mitochondrial-generated ROS and mtDNA lesions and a lower spare respiratory capacity. Silencing of APE1, the major mammalian apurinic/apyrimidinic (AP) endonuclease that participates in the base excision repair (BER) pathway, caused further reductions of spare respiratory capacity in the mutant huntingtin-expressing cells. Localization experiments show that APE1 increases in the mitochondria of wild type Q7 cells but not in the mutant huntingtin Q111 cells after treatment with hydrogen peroxide. Moreover, these results are recapitulated in human HD striata and HD skin fibroblasts that show significant mtDNA damage (increased lesion frequency and mtDNA depletion) and significant decreases in spare respiratory capacity, respectively. These data suggest that mtDNA is a major target of mutant huntingtin-associated oxidative stress and may contribute to subsequent mitochondrial dysfunction and that APE1 (and, by extension, BER) is an important target in the maintenance of mitochondrial function in HD.
mitochondrial DNA; Huntington’s disease; mitochondrial respiration base excision repair; AP endonuclease; mitochondrial dysfunction
We have previously proposed triosephosphate isomerase of Giardia lamblia (GlTIM) as a target for rational drug design against giardiasis, one of the most common parasitic infections in humans. Since the enzyme exists in the parasite and the host, selective inhibition is a major challenge because essential regions that could be considered molecular targets are highly conserved. Previous biochemical evidence showed that chemical modification of the non-conserved non-catalytic cysteine 222 (C222) inactivates specifically GlTIM. The inactivation correlates with the physicochemical properties of the modifying agent: addition of a non-polar, small chemical group at C222 reduces the enzyme activity by one half, whereas negatively charged, large chemical groups cause full inactivation.
In this work we used mutagenesis to extend our understanding of the functional and structural effects triggered by modification of C222. To this end, six GlTIM C222 mutants with side chains having diverse physicochemical characteristics were characterized. We found that the polarity, charge and volume of the side chain in the mutant amino acid differentially alter the activity, the affinity, the stability and the structure of the enzyme. The data show that mutagenesis of C222 mimics the effects of chemical modification. The crystallographic structure of C222D GlTIM shows the disruptive effects of introducing a negative charge at position 222: the mutation perturbs loop 7, a region of the enzyme whose interactions with the catalytic loop 6 are essential for TIM stability, ligand binding and catalysis. The amino acid sequence of TIM in phylogenetic diverse groups indicates that C222 and its surrounding residues are poorly conserved, supporting the proposal that this region is a good target for specific drug design.
The results demonstrate that it is possible to inhibit species-specifically a ubiquitous, structurally highly conserved enzyme by modification of a non-conserved, non-catalytic residue through long-range perturbation of essential regions.
The prevalence of spontaneous mutations increases with age in the male germline; consequently, older men have an increased risk of siring children with genetic disease due to de novo mutations. The lacI transgenic mouse can be used to study paternal age effects, and in this system, the prevalence of de novo mutations increases in the male germline at old ages. Mutagenesis is linked with DNA repair capacity, and base excision repair, which can ameliorate spontaneous DNA damage, decreases in nuclear extracts of spermatogenic cells from old mice. Mice heterozygous for a null allele of the Apex1 gene, which encodes apurinic/apyrimidinic endonuclease I (APEN), an essential base excision repair enzyme, display an accelerated increase in spontaneous germline mutagenesis early in life. Here, the consequences of lifelong reduction of APEN on genetic instability in the male germline were examined, for the first time, at middle and old ages. Mutation frequency increased earlier in spermatogenic cells from Apex1+/− mice (by 6 months of age). Nuclear DNA damage increased with age in the spermatogenic lineage for both wild-type and Apex1+/− mice. By old age, mutation frequencies were similar for wild-type and APEN-deficient mice. Mitochondrial genome repair also depends on APEN, and novel analysis of mitochondrial DNA damage revealed an increase in the Apex1+/− spermatogenic cells by middle age. Thus, Apex1 heterozygosity results in accelerated damage to mitochondrial DNA and spontaneous mutagenesis, consistent with an essential role for APEN in maintaining nuclear and mitochondrial DNA integrity in spermatogenic cells throughout life.
paternal age; mutagenesis; base excision repair; AP endonuclease; lacI; transgenic mice
Many forms of neurodegeneration are associated with oxidative stress and mitochondrial dysfunction. Mitochondria are prominent targets of oxidative damage, however, it is not clear whether mitochondrial DNA (mtDNA) damage and/or its lack of repair are primary events in the delayed onset observed in Huntington’s disease (HD). We hypothesize that an age-dependent increase in mtDNA damage contributes to mitochondrial dysfunction in HD. Two HD mouse models were studied, the 3-nitropropionic acid (3-NPA) chemically induced model and the HD transgenic mice of the R6/2 strain containing 115–150 CAG repeats in the huntingtin gene. The mitochondrial toxin 3-NPA inhibits complex II of the electron transport system and causes neurodegeneration that resembles HD in the striatum of human and experimental animals. We measured nuclear and mtDNA damage by quantitative PCR (QPCR) in striatum of 5- and 24-month-old untreated and 3-NPA treated C57BL/6 mice. Aging caused an increase in damage in both nuclear and mitochondrial genomes. 3-NPA induced 4–6 more damage in mtDNA than nuclear DNA in 5-month-old mice, and this damage was repaired by 48 h in the mtDNA. In 24-month-old mice 3NPA caused equal amounts of nuclear and mitochondrial damage and this damage persistent in both genomes for 48 h. QPCR analysis showed a progressive increase in the levels of mtDNA damage in the striatum and cerebral cortex of 7–12-week-old R6/2 mice. Striatum exhibited eight-fold more damage to the mtDNA compared with a nuclear gene. These data suggest that mtDNA damage is an early biomarker for HD-associated neurodegeneration and supports the hypothesis that mtDNA lesions may contribute to the pathogenesis observed in HD.
Mitochondrial DNA repair; Huntington’s disease; R6/2; 3-Nitropropionic acid
The Saccharomyces cerevisiae APN1 gene that participates in base excision repair has been localized both in the nucleus and the mitochondria. APN1 deficient cells (apn1Δ) show increased mutation frequencies in mitochondrial DNA (mtDNA) suggesting that APN1 is also important for mtDNA stability. To understand APN1-dependent mtDNA repair processes we studied the formation and repair of mtDNA lesions in cells exposed to methyl methanesulfonate (MMS). We show that MMS induces mtDNA damage in a dose-dependent fashion and that deletion of the APN1 gene enhances the susceptibility of mtDNA to MMS. Repair kinetic experiments demonstrate that in wild-type cells (WT) it takes 4 hr to repair the damage induced by 0.1% MMS, whereas in the apn1Δ strain there is a lag in mtDNA repair that results in significant differences in the repair capacity between the two yeast strains. Analysis of lesions in nuclear DNA (nDNA) after treatment with 0.1% MMS shows a significant difference in the amount of nDNA lesions between WT and apn1Δ cells. Interestingly, comparisons between nDNA and mtDNA damage show that nDNA is more sensitive to the effects of MMS treatment. However, both strains are able to repair the nDNA lesions, contrary to mtDNA repair, which is compromised in the apn1Δ mutant strain. Therefore, although nDNA is more sensitive than mtDNA to the effects of MMS, deletion of APN1 has a stronger phenotype in mtDNA repair than in nDNA. These results highlight the prominent role of APN1 in the repair of environmentally induced mtDNA damage.
base excision repair; mitochondrial DNA; alkylating agent
We review the potential impact of DDT on public health in Mexico. DDT production and consumption patterns in Mexico during the last 20 years are described and compared with those in the United States. In spite of the restrictions on DDT use in antimalaria campaigns in Mexico, use of DDT is still higher than in other Latin American countries. We analyzed information from published studies to determine accumulated levels of this insecticide in blood, adipose tissue, and breast milk samples from Mexican women. Current lipid-adjusted DDE levels from women living in Mexico City are 6.66 ppb in mammary adipose tissue and 0.594 ppm in total breast milk. Finally, the methodological limitations of existing epidemiological studies on DDT exposure and breast cancer are discussed. We conclude that DDT use in Mexico is a public health problem, and suggest two solutions: identification of alternatives for the control of malaria and educational intervention to reduce DDT exposure. We also recommend strengthening epidemiological studies to evaluate the association between accumulated DDT levels in adipose tissue and breast cancer incidence among Mexican women.
Coronavirus (CoV) envelope (E) protein ion channel activity was determined in channels formed in planar lipid bilayers by peptides representing either the transmembrane domain of severe acute respiratory syndrome CoV (SARS-CoV) E protein, or the full-length E protein. Both of them formed a voltage independent ion conductive pore with symmetric ion transport properties. Mutations N15A and V25F located in the transmembrane domain, prevented the ion conductivity. E protein derived channels showed no cation preference in non-charged lipid membranes, whereas they behaved as pores with mild cation selectivity in negatively-charged lipid membranes. The ion conductance was also controlled by the lipid composition of the membrane. Lipid charge also regulated the selectivity of a HCoV-229E E protein derived peptide. These results suggested that the lipids are functionally involved in E protein ion channel activity, forming a protein-lipid pore, a novel concept for CoV E protein ion channel entity.
Coronavirus; SARS; envelope protein; ion channel; HCoV-229E; lipid membranes
The regioselectivity of the epoxide ring opening of 2-methyl-3,4-epoxy alcohols with diethylpropynylalane has been studied as a function of the C1 alcohol protecting group. An efficient selective method was developed using MEM as the protecting group. The reaction proceeded in a highly regioselective manner providing the useful 1,3-diol motif. The undesired 1,4-diol product produced by some free alcohol diastereomers was not observed. This highly stereoselective method provides access to termini-differentiated stereotetrads, which are essential building bocks for polypropionate synthesis.
cleavage; epoxides; aluminum; stereotetrads; 3, 4-epoxy alcohols
In agreement with historical documentation, several genetic studies have revealed ancestral links between the European Romani and India. The entire mitochondrial DNA (mtDNA) of 27 Spanish Romani was sequenced in order to shed further light on the origins of this population. The data were analyzed together with a large published dataset (mainly hypervariable region I [HVS-I] haplotypes) of Romani (N = 1,353) and non-Romani worldwide populations (N>150,000). Analysis of mitogenomes allowed the characterization of various Romani-specific clades. M5a1b1a1 is the most distinctive European Romani haplogroup; it is present in all Romani groups at variable frequencies (with only sporadic findings in non-Romani) and represents 18% of their mtDNA pool. Its phylogeographic features indicate that M5a1b1a1 originated 1.5 thousand years ago (kya; 95% CI: 1.3–1.8) in a proto-Romani population living in Northwest India. U3 represents the most characteristic Romani haplogroup of European/Near Eastern origin (12.4%); it appears at dissimilar frequencies across the continent (Iberia: ∼31%; Eastern/Central Europe: ∼13%). All U3 mitogenomes of our Iberian Romani sample fall within a new sub-clade, U3b1c, which can be dated to 0.5 kya (95% CI: 0.3–0.7); therefore, signaling a lower bound for the founder event that followed admixture in Europe/Near East. Other minor European/Near Eastern haplogroups (e.g. H24, H88a) were also assimilated into the Romani by introgression with neighboring populations during their diaspora into Europe; yet some show a differentiation from the phylogenetically closest non-Romani counterpart. The phylogeny of Romani mitogenomes shows clear signatures of low effective population sizes and founder effects. Overall, these results are in good agreement with historical documentation, suggesting that cultural identity and relative isolation have allowed the Romani to preserve a distinctive mtDNA heritage, with some features linking them unequivocally to their ancestral Indian homeland.
Background: The distribution of polymorphisms in the CYP2D6 and CYP2C19 genes allows inferring the potential risk for specific adverse drug reactions and lack of therapeutic effects in humans. This variability shows differences among human populations. The aim of this study was to analyze single-nucleotide polymorphisms related to a poor metabolizer (PM) phenotype in nonpreviously studied Amerindian groups and Mestizos (general admixed population) from Mexico. Methods: We detected by SNaPshot® different polymorphisms located in CYP2D6 (*3, *4, *6, *7, and *8) and CYP2C19 (*2, *3, *4 and *5) in western Mestizos (n=145) and five Amerindian groups from Mexico: Tarahumaras from the North (n=88); Purépechas from the Center (n=101); and Tojolabales (n=68), Tzotziles (n=88), and Tzeltales (n=20) from the Southeast. Genotypes were observed by capillary electrophoresis. The genetic relationships among these populations were estimated based on these genes. Results and Discussion: The wild-type allele (*1) of both genes was predominant in the Mexican populations studied. The most widely observed alleles were CYP2C19*2 (range, 0%–31%) and CYP2D6*4 (range, 1.2%–7.3%), whereas CYP2D6*3 was exclusively detected in Mestizos. Conversely, CYP2C19*4 and *5, as well as CYP2D6*3, *6, *7, and *8, were not observed in the majority of the Mexican populations. The Tarahumaras presented a high frequency of the allele CYP2C19*2 (31%) and of homozygotes *2/*2 (10.7%), which represent a high frequency of potentially PM phenotypes in this Amerindian group. The genetic distances showed high differentiation of Tarahumaras (principally for CYP2C19 gene). In general, a relative proximity was observed between most of the Amerindian, Mexican-Mestizo, and Latin-American populations. Conclusion: In general, the wild-type allele (*1) predominates in Mexican populations, outlining a relatively homogeneous distribution for CYP2C19 and CYP2D6. The exception is the Tarahumara group that displays a potentially increased risk for adverse reactions to CYP2C19-metabolized drugs.
Various hepatoprotective herbal products from plants are available in Mexico, where up to 85% of patients with liver disease use some form of complementary and alternative medicine. However, only few studies have reported on the biological evaluation of these products.
Using a model of carbon tetrachloride (CCl4)-induced hepatotoxicity in rats, we evaluated the effects of commercial herbal extracts used most commonly in the metropolitan area of Monterrey, Mexico.
Materials and Methods:
The commercial products were identified through surveys in public areas. The effect of these products given with or without CCl4 in rats was evaluated by measuring the serum concentrations of aspartate amino transferase (AST) and alanine amino transferase (ALT), and histopathological analysis. Legalon® was used as the standard drug.
The most commonly used herbal products were Hepatisan® capsules, Boldo capsules, Hepavida® capsules, Boldo infusion, and milk thistle herbal supplement (80% silymarin). None of the products tested was hepatotoxic according to transaminase and histological analyses. AST and ALT activities were significantly lower in the Hepavida+CCl4-treated group as compared with the CCl4-only group. AST and ALT activities in the silymarin, Hepatisan, and Boldo tea groups were similar to those in the CCl4 group. The CCl4 group displayed submassive confluent necrosis and mixed inflammatory infiltration. Both the Hepatisan+CCl4 and Boldo tea+CCl4 groups exhibited ballooning degeneration, inflammatory infiltration, and lytic necrosis. The silymarin+CCl4 group exhibited microvesicular steatosis. The Hepavida+CCl4- and Legalon+CCL4-treated groups had lower percentages of necrotic cells as compared with the CCl4-treated group; this treatment was hepatoprotective against necrosis.
Only Hepavida had a hepatoprotective effect.
Alanine transferase; aspartate transferase; hepatoprotection; liver injury; natural products
In most clinical trials, human mesenchymal stem cells (hMSCs) are expanded in vitro before implantation. The genetic stability of human stem cells is critical for their clinical use. However, the relationship between stem-cell expansion and genetic stability is poorly understood. Here, we demonstrate that within the normal expansion period, hMSC cultures show a high percentage of aneuploid cells that progressively increases until senescence. Despite this accumulation, we show that in a heterogeneous culture the senescence-prone hMSC subpopulation has a lower proliferation potential and a higher incidence of aneuploidy than the non-senescent subpopulation. We further show that senescence is linked to a novel transcriptional signature that includes a set of genes implicated in ploidy control. Overexpression of the telomerase catalytic subunit (human telomerase reverse transcriptase, hTERT) inhibited senescence, markedly reducing the levels of aneuploidy and preventing the dysregulation of ploidy-controlling genes. hMSC-replicative senescence was accompanied by an increase in oxygen consumption rate (OCR) and oxidative stress, but in long-term cultures that overexpress hTERT, these parameters were maintained at basal levels, comparable to unmodified hMSCs at initial passages. We therefore propose that hTERT contributes to genetic stability through its classical telomere maintenance function and also by reducing the levels of oxidative stress, possibly, by controlling mitochondrial physiology. Finally, we propose that aneuploidy is a relevant factor in the induction of senescence and should be assessed in hMSCs before their clinical use.
Stem cells; MSC; genetic instability; telomerase; aneuploidy; mitochondrial metabolism
It has been suggested that there is an inverse association between breastfeeding and the risk of childhood cancer. We investigated the association between full breastfeeding and paediatric cancer (PC) in a case control study in Spain.
Maternal reports of full breastfeeding, collected through personal interviews using the Paediatric Environmental History, were compared among 187 children 6 months of age or older who had PC and 187 age-matched control siblings.
The mean duration of full breastfeeding for cases were 8.43 and 11.25 weeks for controls. Cases had been significantly more often bottle-fed than controls (odds ratio (OR) 1.8; 95% confidence interval (CI) 1.1–2.8). Cases were significantly less breastfed for at least 2 months (OR 0.5; 95% CI 0.3–0.8), for at least 4 months (OR 0.5; 95% CI 0.3–0.8), and for 24 weeks or more (OR 0.5; 95% CI 0.2–0.9).
Breastfeeding was inversely associated with PC, the protection increasing with the duration of full breastfeeding. Additional research on possible mechanisms of this association may be warranted. Meanwhile, breastfeeding should be encouraged among mothers.
full breastfeeding; paediatric oncology; risk factor
Human tuberculosis caused by M. bovis is a zoonosis presently considered sporadic in developed countries, but remains a poorly studied problem in low and middle resource countries. The disease in humans is mainly attributed to unpasteurized dairy products consumption. However, transmission due to exposure of humans to infected animals has been also recognized. The prevalence of tuberculosis infection and associated risk factors have been insufficiently characterized among dairy farm workers (DFW) exposed in settings with poor control of bovine tuberculosis.
Tuberculin skin test (TST) and Interferon-gamma release assay (IGRA) were administered to 311 dairy farm and abattoir workers and their household contacts linked to a dairy production and livestock facility in Mexico. Sputa of individuals with respiratory symptoms and samples from routine cattle necropsies were cultured for M. bovis and resulting spoligotypes were compared. The overall prevalence of latent tuberculosis infection (LTBI) was 76.2% (95% CI, 71.4–80.9%) by TST and 58.5% (95% CI, 53.0–64.0%) by IGRA. Occupational exposure was associated to TST (OR 2.72; 95% CI, 1.31–5.64) and IGRA (OR 2.38; 95% CI, 1.31–4.30) adjusting for relevant variables. Two subjects were diagnosed with pulmonary tuberculosis, both caused by M. bovis. In one case, the spoligotype was identical to a strain isolated from bovines.
We documented a high prevalence of latent and pulmonary TB among workers exposed to cattle infected with M. bovis, and increased risk among those occupationally exposed in non-ventilated spaces. Interspecies transmission is frequent and represents an occupational hazard in this setting.
Mycobacterium tuberculosis complex causes tuberculosis in humans and other mammals. The complex includes M. bovis, which causes bovine tuberculosis. The main route of transmission of this zoonosis is the consumption of unpasteurized dairy products. Nevertheless, exposure to infected cattle while performing husbandry and farm activities may cause disease as well. In this study we were able to demonstrate: 1) A high prevalence of tuberculosis asymptomatic infection (latent tuberculosis) among workers exposed to infected cattle; 2) A higher probability of infection among individuals who are occupationally exposed in closed spaces; and 3) Cattle to human transmission confirmed by molecular methods (spoligotyping). We conclude that occupational exposure is frequent, and therefore strict prevention and control measures are required in these settings.
Background. Lipoprotein(a) [Lp(a)] is a known risk factor for cardiovascular disease, yet its influence on metabolic syndrome (MS) is still controversial. The purpose of this study was to assess the impact generated by this diagnosis in serum Lp(a) concentrations. Materials and Methods. A total of 1807 subjects of both genders (55.3% women and 44.7% men) belonging to the Maracaibo City Metabolic Syndrome Prevalence Study were evaluated. Results were expressed as Mean ± SD, determining differences through Student's t-test and One-Way ANOVA test. Multiple logistic regression models were utilized for analyzing factors associated with elevated serum Lp(a) levels and MS. Total cholesterol and LDL-C were corrected according to Lp(a)-Cholesterol when necessary.
Results. No differences were found in Lp(a) values between genders; P = 0,292. The association between MS and the classification of Lp(a) was statistically significant (χ2 = 28.33; P < 0,0001), with greater levels in subjects with this diagnosis. In the univariate analysis, subjects with each of the separate diagnostic criteria showed higher serum Lp(a) concentrations, except for hyperglycemia. Conclusions. Lp(a) values exhibit important variations regarding MS and each of its components. Impaired fasting glucose appeared as a protecting factor against elevated Lp(a) concentrations, whereas its association with LDL-C and hs-CRP suggests a potential pro-inflammatory role.
Uptake through the Dopamine Transporter (DAT) is the primary mechanism of terminating dopamine signaling within the brain, thus playing an essential role in neuronal homeostasis. Deregulation of DAT function has been linked to several neurological and psychiatric disorders including ADHD, schizophrenia, Parkinson’s disease, and drug addiction. Over the last 15 years, several studies have revealed a plethora of mechanisms influencing the activity and cellular distribution of DAT; suggesting that fine-tuning of dopamine homeostasis occurs via an elaborate interplay of multiple pathways. Here, we show for the first time that the βγ subunits of G proteins regulate DAT activity. In heterologous cells and brain tissue, a physical association between Gβγ subunits and DAT was demonstrated by co-immunoprecipitation. Furthermore, in vitro pull-down assays using purified proteins established that this association occurs via a direct interaction between the intracellular carboxy-terminus of DAT and Gβγ. Functional assays performed in the presence of the non-hydrolyzable GTP analog GTP-γ-S, Gβγ subunit overexpression, or the Gβγ activator mSIRK all resulted in rapid inhibition of DAT activity in heterologous systems. Gβγ activation by mSIRK also inhibited dopamine uptake in brain synaptosomes and dopamine clearance from mouse striatum as measured by high-speed chronoamperometry in vivo. Gβγ subunits are intracellular signaling molecules that regulate a multitude of physiological processes through interactions with enzymes and ion channels. Our findings add neurotransmitter transporters to the growing list of molecules regulated by G-proteins and suggest a novel role for Gβγ signaling in the control of dopamine homeostasis.
A prospective, non-randomised, transversal and comparative study, carried out in INOVA Vision Institute and Autonomous University of Aguascalientes. Pterygium is an important illness that affects 22% people from tropic and equatorial zones. Is an inflammatory process caused by UV rays, and it has a behavior similar to a neoplasm. For this study was taken into consideration 191 samples from the INOVA Vision Institute, Aguascalientes, Mexico. Include 73 pterygia samples, which were obtained during resection under sterile conditions. 44 normal conjunctiva samples were obtained from the same patients when harvesting the conjunctival autograft, or from other patients undergoing extracapsular cataract extraction from the superior bulbar region. Tears from patients with pterygium (n = 50) and normal volunteers (n = 24) were obtained using a calibrated glass micro capillary tube. The surgical conjunctiva and pterygia samples were subjected to reverse-transcription polymerase chain reaction (RT-PCR), western blot, and immunohistochemistry. Tears were analyzed by enzyme-linked immunosorbent assays.
This was a prospective, non-randomised study involving 191 biological samples taken from patients with pterygium and normal volunteers, whom were operated under local anaesthesia by either complete resection of the lesion with primary closure, or resection with conjunctival autograft. Tissue samples were fixed in 10% formaldehyde. Sections were routinely stained with hematoxylin and eosin. HCC expression was evaluated by reverse-transcription polymerase chain reaction (RT-PCR), immunohistochemistry, and by western blotting. All tears samples were analyzed by enzyme-linked immunosorbent assays (ELISA).
Expression levels and distribution patterns of HCC in normal conjunctiva and pterygium. Higher levels of HCC mRNAs and proteins were detected in pterygium compared with a normal conjunctiva. Immunohistochemistry revealed that HCC was localized in the apical cells of the epithelium in the normal conjunctiva. In contrast, HCC was detected in all extension of epithelial tissue, from apical to basal cells in pterygia. The concentration of HCC protein in tears was higher in patients with pterygium versus controls.
HCC may play an important role in protecting normal conjunctiva, and regulating inflammatory conditions of the anterior ocular surface.
Pterygium; Human cystatin C; Proliferation
Cannabinoids, the active components of marijuana and their derivatives, are currently investigated due to their potential therapeutic application for the management of many different diseases, including cancer. Specifically, Δ9-Tetrahydrocannabinol (THC) and Cannabidiol (CBD) – the two major ingredients of marijuana – have been shown to inhibit tumor growth in a number of animal models of cancer, including glioma. Although there are several pharmaceutical preparations that permit the oral administration of THC or its analogue nabilone or the oromucosal delivery of a THC- and CBD-enriched cannabis extract, the systemic administration of cannabinoids has several limitations in part derived from the high lipophilicity exhibited by these compounds. In this work we analyzed CBD- and THC-loaded poly-ε-caprolactone microparticles as an alternative delivery system for long-term cannabinoid administration in a murine xenograft model of glioma. In vitro characterization of THC- and CBD-loaded microparticles showed that this method of microencapsulation facilitates a sustained release of the two cannabinoids for several days. Local administration of THC-, CBD- or a mixture (1∶1 w:w) of THC- and CBD-loaded microparticles every 5 days to mice bearing glioma xenografts reduced tumour growth with the same efficacy than a daily local administration of the equivalent amount of those cannabinoids in solution. Moreover, treatment with cannabinoid-loaded microparticles enhanced apoptosis and decreased cell proliferation and angiogenesis in these tumours. Our findings support that THC- and CBD-loaded microparticles could be used as an alternative method of cannabinoid delivery in anticancer therapies.
Knowledge on the patterns of repetition amongst individuals who develop language deficits in association with right hemisphere lesions (crossed aphasia) is very limited. Available data indicate that repetition in some crossed aphasics experiencing phonological processing deficits is not heavily influenced by lexical-semantic variables (lexicality, imageability, and frequency) as is regularly reported in phonologically-impaired cases with left hemisphere damage. Moreover, in view of the fact that crossed aphasia is rare, information on the role of right cortical areas and white matter tracts underpinning language repetition deficits is scarce. In this study, repetition performance was assessed in two patients with crossed conduction aphasia and striatal/capsular vascular lesions encompassing the right arcuate fasciculus (AF) and inferior frontal-occipital fasciculus (IFOF), the temporal stem and the white matter underneath the supramarginal gyrus. Both patients showed lexicality effects repeating better words than non-words, but manipulation of other lexical-semantic variables exerted less influence on repetition performance. Imageability and frequency effects, production of meaning-based paraphrases during sentence repetition, or better performance on repeating novel sentences than overlearned clichés were hardly ever observed in these two patients. In one patient, diffusion tensor imaging disclosed damage to the right long direct segment of the AF and IFOF with relative sparing of the anterior indirect and posterior segments of the AF, together with fully developed left perisylvian white matter pathways. These findings suggest that striatal/capsular lesions extending into the right AF and IFOF in some individuals with right hemisphere language dominance are associated with atypical repetition patterns which might reflect reduced interactions between phonological and lexical-semantic processes.
right hemisphere; language; crossed aphasia; conduction aphasia; language network; structural connectivity
Clinical Practice Guidelines recommend using peripheral blood pulse measuring as a screening test for Atrial Fibrillation. However, there is no adequate evidence supporting the efficacy of such procedure in primary care clinical practice. This paper describes a study protocol designed to verify whether early opportunistic screening for Atrial Fibrillation by measuring blood pulse is more effective than regular practice in subjects aged 65 years attending primary care centers.
An cluster-randomized controlled trial conducted in Primary Care Centers of the Spanish National Health Service. A total of 269 physicians and nurses will be allocated to one of the two arms of the trial by stratified randomization with a 3:2 ratio (three practitioners will be assigned to the Control Group for every two practitioners assigned to the Experimental Group). As many as 12 870 patients aged 65 years or older and meeting eligibility criteria will be recruited (8 580 will be allocated to the Experimental Group and 4 290 to the Control Group). Randomization and allocation to trial groups will be carried out by a central computer system. The Experimental Group practitioners will conduct an opportunistic case finding for patients with Atrial Fibrillation, while the Control Group practitioners will follow the regular guidelines. The first step will be finding new Atrial Fibrillation cases. A descriptive inferential analysis will be performed (bivariate and multivariate by multilevel logistic regression analysis).
If our hypothesis is confirmed, we expect Primary Care professionals to take a more proactive approach and adopt a new protocol when a patient meeting the established screening criteria is identified. Finally, we expect this measure to be incorporated into Clinical Practice Guidelines.
The study is registered as NCT01291953 (ClinicalTrials.gob)
Atrial fibrillation; Screening; Opportunistic case finding; Secondary prevention; Primary care
Obesity has become a global epidemic and a leading metabolic disease in the world. Laparoscopic surgeries may influence the function of the immunologic system. The percentages of CD4+ and CD8+ T lymphocyte cells have been described as prognostic factors for patients undergoing abdominal surgeries. This study aimed to evaluate the changes in CD4+ and CD8+ T lymphocyte cells, the ratio of CD4+ to CD8+ cells, and the ZAP-70 kinase expression on T CD3+ and B CD19+ cells in obese and normal-weight individuals undergoing laparoscopic cholecystectomy (LC).
The study group consisted of 46 asymptomatic patients with gallstones shown by ultrasound examination but without signs of any gallbladder complications. The patients underwent planned LC. Blood samples were obtained at three times, and the percentages of studied cells were measured by flow cytometry. Patients were enrolled to two groups: N group (body mass index [BMI], ≤25 kg/m2) and O group (BMI, ≥30 kg/m2). For statistical analysis, the Mann–Whitney U test and the Wilcoxon matched-pairs signed-ranks test were used. All p values lower than 0.05 were considered significant.
The percentage of CD4+ T cells did not differ between the N and O groups before or after the surgery. Only in the N group did the percentage of CD4+ lymphocytes increase from 0 to 48 h. A higher percentage of CD8+ lymphocytes was observed in the O group postoperatively than in the N group. Differences of ZAP-70 kinase expression in the O group were observed at 24 and 48 h of the study. Decreased expression of ZAP-70 kinase was shown in the N group at both 0–24 and 24–48 h. In the O group, this tendency was noted at 24–48 h.
Immunologic activation after LC was confirmed in both weight groups. However, higher modulation, more typical for open surgeries, was observed in the obese group.
Laparoscopy; Obesity; Pneumoperitoneum; T cells; ZAP-70 kinase; CD4+; CD8+
Shear stress stimulates nitric oxide (NO) release in endothelial cells. Stretch-activated ion channels (SACs) and the transient receptor potential vanilloid type 1 (TRPV1) receptor respond to mechanical stimulus and are permeable to Na+, Ca2+ and K+. The influence of SACs and the TRPV1 receptor on NO release on the heart and on the vascular reactivity of the thoracic aorta (TA) was studied. Experiments were performed in isolated perfused heart, cultured endothelial cells and TA rings from Wistar rats. Capsaicin (10 μM, 30 μM) was used as a NO release stimulator, capsazepine (6 μM, 10 μM) was used as a capsaicin antagonist and gadolinium (3 μM, 5 μM) was used as an inhibitor of SACs. NO was measured by the Kelm and Tenorio methods. Left ventricular pressure was recorded and coronary vascular resistance was calculated. Capsaicin increased NO release in the heart by 58% (395±8 pmol/mL to 627±23 pmol/mL). Capsazepine and gadolinium inhibited NO release by 74% and 82%, respectively. This tendency was similar in all experimental models. Capsaicin attenuated the effects of norepinephrine (10 M to 7 M) on TA and had no effect in the presence of Nω-nitro-L-arginine methyl ester. Therefore, the authors conclude that SACs and the TRPV1 receptor are both present in the coronary endothelium and that both participate in Ca2+-dependent NO release.
Endothelium; Nitric oxide; Stretch-activated ion channels; TRPV1 receptors
Alterations of mTOR gene expression have been implicated in the pathogenesis of endometrioid endometrial cancer however only few studies explored the cause of increased mTOR activation in this malignancy. miRNAs are small, noncoding RNAs, which were proven to regulated gene expression at the posttranscriptional level. The study aimed to explore deregulation of miRNAs targeting mTOR kinase (miR-99a, miR-100 and miR-199b) as a possible cause of its altered expression in EEC tissues. In addition expression of the three miRNAs was investigated in plasma of EEC patients and was assessed in terms of diagnostic and prognostic utility.
We investigated expression of mTOR kinase transcripts in 46 fresh tissue samples. Expression of miR-99a, miR-100 and miR-199b was investigated in the same group of fresh samples, and in additional 58 FFPE sections as well as in 48 plasma samples using qPCR. Relative quantification was performed using experimentally validated endogenous controls.
mTOR kinase expression was increased in EEC tissues and was accompanied by decreased expression of all three miRNAs. Down-regulation of the investigated miRNAs was discovered in plasma of EEC patients and miRNA signatures classified EEC tissues (miR-99a/miR-100/miR-199b) and plasma (miR-99a/miR-199b) samples with higher accuracy in comparison to single miRNAs. We also revealed that miR-100 was an independent prognostic marker of overall survival.
We conclude that increased expression of mTOR kinase coexists with down-regulation of its targeting miRNAs, which could suggest a new mechanism of mTOR pathway alterations in EEC. In addition, our findings implicate that miRNA signatures can be considered promising biomarkers for early detection and prognosis of endometrioid endometrial carcinoma.
mTOR; microRNA; miRNA; Endometrial cancer; Plasma
Trained people exhibit low plasma concentrations of triacylglcyerols in both fasting and postprandial states. Exercise practice is commonly believed to improve postprandial lipemia. In addition, elevated postprandial lipemia is an indicator of poor lipid clearance, and it has been associated with atherosclerosis, insulin resistance, and obesity. Spirulina maxima is an edible microorganism with a high nutritional value. When it is consumed, beneficial properties to health have been demonstrated, such as hypolipemic and antihypertensive properties in human beings. This work evaluates the effects of orally administrated S. maxima on postprandial lipemia in a young Mexican sporting population after 15 days of consumption, as a possible alternative treatment to improve their lipid clearance. Forty-one runners (10–26 years old; 21 men and 20 women) volunteered to participate in the study. All of them were physically active for at least 1 year before the study and were not undergoing training during the study. The subjects consumed 5 g of Spirulina during 15 days. Before and after the treatment with Spirulina, they consumed (12 h fasting) a standardized meal with high fat content (53.2% total calories). Postprandial lipemia was measured at 1.5, 3, and 4.5 h after the fatty meal. Fasting plasma triacylglycerol (TAG) concentrations were lower after Spirulina treatment than before treatment. In addition, the postprandial area under the curve of TAG concentrations was lower after the treatment with Spirulina. Sixty-two percent of the youngest runners (10–16 years) studied exhibited the best response to the treatment. Orally administered S. maxima decreased postprandial lipemia in sporting teenagers. The youngest people were the most responsive to the beneficial effects of Spirulina on postprandial lipemia.
area under curve; exercise; nutrition; triacylglycerols
Introduction and Objectives
Lymphatic metastasis is a common occurrence in human breast cancer, mechanisms remaining poorly understood. MDA-MB-468LN (468LN), a variant of the MDA-MB-468GFP (468GFP) human breast cancer cell line, produces extensive lymphatic metastasis in nude mice. 468LN cells differentially express α9β1 integrin, a receptor for lymphangiogenic factors VEGF-C/-D. We explored whether (1) differential production of VEGF-C/-D by 468LN cells provides an autocrine stimulus for cellular motility by interacting with α9β1 and a paracrine stimulus for lymphangiogenesis in vitro as measured with capillary-like tube formation by human lymphatic endothelial cells (HMVEC-dLy); (2) differential expression of α9 also promotes cellular motility/invasiveness by interacting with macrophage derived factors; (3) stable knock-down of VEGF-D or α9 in 468LN cells abrogates lymphangiogenesis and lymphatic metastasis in vivo in nude mice.
A comparison of expression of cyclo-oxygenase (COX)-2 (a VEGF-C/-D inducer), VEGF-C/-D and their receptors revealed little COX-2 expression by either cells. However, 468LN cells showed differential VEGF-D and α9β1 expression, VEGF-D secretion, proliferative, migratory/invasive capacities, latter functions being stimulated further with VEGF-D. The requirement of α9β1 for native and VEGF-D-stimulated proliferation, migration and Erk activation was demonstrated by treating with α9β1 blocking antibody or knock-down of α9. An autocrine role of VEGF-D in migration was shown by its impairment by silencing VEGF-D and restoration with VEGF-D. 468LN cells and their soluble products stimulated tube formation, migration/invasiveness of HMVEC-dLy cell in a VEGF-D dependent manner as indicated by the loss of stimulation by silencing VEGF-D in 468LN cells. Furthermore, 468LN cells showed α9-dependent stimulation of migration/invasiveness by macrophage products. Finally, capacity for intra-tumoral lymphangiogenesis and lymphatic metastasis in nude mice was completely abrogated by stable knock-down of either VEGF-D or α9 in 468LN cells.
Differential capacity for VEGF-D production and α9β1 integrin expression by 468LN cells jointly contributed to their lymphatic metastatic phenotype.