Sulphur Amino Acids: How Important is it to Set Reference Ranges for Each Population?

INTRODUCTION: Metabolism of sulfur amino acids requires an optimal interplay between nutritional demand, enzymes, transporters, and adequate dietary intake of B vitamins. Insufficient intake and excess are detrimental, and concentrations depend on health status. However, plasma aminothiol concentrations, previously reported in healthy subjects using highly sensitive methods, vary considerably, and age- and gender differences were observed. Therefore, defining age- and gender-specific ranges for each population is crucial to evaluate the meaning of plasma thiol redox state in health and disease. METHODS: A healthy Portuguese pediatric population (n=90), aged 9- (n=38) and 17-year-old (n=52), was evaluated. Plasma aminothiols, total homocysteine (tHcy), cysteine (tCys), glutathione (tGSH) and γ-glutamylcysteine (tγ-Glu-Cys), were analysed as SBD-F derivatives by HPLC with fluorescence detection. RESULTS/CASE REPORT: Mean plasma concentrations (SD) for the 9- and the 17-year-old groups, were as following: tHcy = 4.58 (0.98); 8.13 (3.27) µM, p <0.001; tCys = 207.34 (32.07); 198.59 (21.24) µM, p = 0.274; tGSH = 4.54 (1.08); 5.20 (1.84) µM, p = 0.123 and tγ-Glu-Cys = 1.47 (0.30); 1.06 (0.28) µM, p < 0.001, respectively. No statistically significant differences were found between males and females in the 9-year-old group. However, in the 17-year-old group, significant differences between genders were observed for tHcys (p < 0.001) and tγ-Glu-Cys (p = 0.039), with males presenting the highest concentrations. When correlating the four thiols' plasma concentrations, only the precursors of glutathione, tγ-Glu-Cys and tCys, were positively correlated (r = 0.450, p < 0.001). CONCLUSION: Our results showed significant differences in tHcy and tγ-Glu-Cys levels across both age groups, which increased and decreased with age, respectively. It is interesting to highlight that in the 17-year-old group, tHcy and tγ-Glu-Cys levels were higher in males than in females. These observations showed that age and gender influence plasma levels of thiols, which may impact cellular oxidative status. In conclusion, setting age and gender distinct ranges for each specific population is of utmost importance for understanding disease mechanisms and the effectiveness of therapeutic interventions.

Suppressing Aneuploidy-Associated Phenotypes Improves the Fitness of Trisomy 21 Cells.

An abnormal number of chromosomes, or aneuploidy, accounts for most spontaneous abortions, causes developmental defects, and is associated with aging and cancer. The molecular mechanisms by which aneuploidy disrupts cellular function remain largely unknown. Here, we show that aneuploidy disrupts the morphology of the nucleus. Mutations that increase the levels of long-chain bases suppress nuclear abnormalities of aneuploid yeast independent of karyotype identity. Quantitative lipidomics indicates that long-chain bases are integral components of the nuclear membrane in yeast. Cells isolated from patients with Down syndrome also show that abnormal nuclear morphologies and increases in long-chain bases not only suppress these abnormalities but also improve their fitness. We obtained similar results with cells isolated from patients with Patau or Edward syndrome, indicating that increases in long-chain bases improve the fitness of aneuploid cells in yeast and humans. Targeting lipid biosynthesis pathways represents an important strategy to suppress nuclear abnormalities in aneuploidy-associated diseases.

La especialidad médica de gestión en salud en la Universidad Nacional Mayor de San Marcos: 30 años formando gestores para el sector salud / The medical major of health management in Universidad Nacional Mayor de San Marcos: 30 years training managers for the health sector

Se presenta un recuento histórico de los 30 años de la especialidad médica de Gestión en Salud en la Universidad Nacional Mayor de San Marcos. La especialidad fue creada en 1988 como Medicina General Integral en respuesta a la escasez de médicos especialistas en gestión. El nombre de la especialidad se mantuvo hasta 1994, cuando cambia a Medicina Integral y Gestión en Salud para enfatizar su orientación gerencial. En 2002, se modifica el plan curricular considerando la importancia de tener una especialización médica exclusiva de gestión para el sector salud. La especialidad es pionera en una formación gradual en tres niveles: microgestión, mesogestión y macrogestión. El nuevo plan curricular permitió a los residentes la posibilidad de mejores rotaciones en instituciones públicas y privadas. A partir del 2007, la especialidad es denominada Gestión en Salud, cuyos médicos especialistas mantienen su sólida formación para desempeñarse en los procesos de decisión, conducción, dirección y operatividad de los sistemas de salud. La visión de los creadores de la especialidad se encuentra vigente, los especialistas de Gestión en Salud se desempeñan en las diferentes instituciones del sistema de salud, aportando sus conocimientos y habilidades, y generando un impacto en la salud de la población peruana.

A historical account of the 30 years of the medical major of Health Management at Universidad Nacional Mayor de San Marcos is presented. The major was created in 1988 as Comprehensive General Medicine in response to the shortage of management specialists. The name of the major remained until 1994, when it changed to Integral Medicine and Health Management to emphasize its managerial orientation. In 2002, the curricular plan was modified considering the importance of having an exclusive medical specialization in management for the health sector. The specialty is a pioneer in a gradual education in three levels: micro-management, meso-management, and macro-management. The new curricular plan allowed residents the possibility to access better rotations at public and private institutions. Since 2007, the major is labeled Health Management, and its specialists maintain their solid training that enables them to perform in the decision-making, management, administration, and operation processes of the health systems. The vision of the creators of this major is current to this day; Health Management specialists work in the different institutions of the health system, contributing their knowledge and skills, and generating an impact on the health of the Peruvian population.

Scedosporium apiospermum and S. prolificans mixed disseminated infection in a lung transplant recipient: An unusual case of long-term survival with combined systemic and local antifungal therapy in intensive care unit.

Infections due Scedosporium spp. in lung transplant recipients are associated with disseminated disease with high mortality rates. The adjunctive local antifungal therapy may be a useful option when systemic treatment is insufficient and/or surgery is not feasible. We present a case of mixed disseminated infection due Scedosporium apiospermum and S. prolificans in a lung transplant recipient. Combined local and systemic antifungal therapy provided an unusual long-term survival in the intensive care unit.

Changes in macrophage function modulated by the lipid environment.

Macrophages (Mφs) play a critical role in the defense against pathogens, orchestrating the inflammatory response during injury and maintaining tissue homeostasis. During these processes, macrophages encounter a variety of environmental conditions that are likely to change their gene expression pattern, which modulates their function. In this study, we found that murine Mφs displayed two different subpopulations characterized by differences in morphologies, expression of surface markers and phagocytic capacity under non-stimulated conditions. These two subpopulations could be recapitulated by changes in the culture conditions. Thus, Mφs grown in suspension in the presence of serum were highly phagocytic, whereas subtraction of serum resulted in rapid attachment and reduced phagocytic activity. The difference in phagocytosis between these subpopulations was correlated with the expression levels of FcγR. These two cell subpopulations also differed in their responses to LPS and the expression of surface markers, including CD14, CD86, scavenger receptor A1, TLR4 and low-density lipoprotein receptor. Moreover, we found that the lipid/cholesterol content in the culture medium mediated the differences between these two cell subpopulations. Thus, we described a mechanism that modulates Mφ function depending on the exposure to lipids within their surrounding microenvironment.

Small aminothiol compounds improve the function of Arg to Cys variant proteins: effect on the human cystathionine ß-synthase p.R336C.

The key regulatory point of L-methionine (Met) and L-homocysteine (Hcy) degradation is catalyzed by cystathionine beta-synthase (CBS). CBS deficiency is caused by mutations in CBS gene, often resulting in protein misfolding. The prevalence of CBS deficiency in Qatar is 1/1800, ∼200-fold higher than the worldwide prevalence of 1/344 000. Almost all patients bear the CBS p.R336C variant. More than 20 years ago, it was shown in vitro that two unrelated protein variants with a substitution of an arginine (Arg) residue by cysteine (Cys) could be rescued by cysteamine (mercaptoethylamine), likely via formation of a disulfide between Cys and cysteamine, functionally mimicking the wild-type (WT) Arg side-chain. Based on these findings, we aimed to study whether cysteamine was able to improve the function of p.R336C CBS variant. Additionally, we tested the effect of mercaptoethylguanidine (MEG), a compound with a guanidino and a thiol function that may resemble Arg structure better than cysteamine. Three purified recombinant CBS proteins (p.R336C, p.R336H and WT) were pre-incubated with cysteamine, MEG or Cys (as negative control), and CBS activity and stability were measured. Pre-incubation with cysteamine and MEG increased the enzymatic activity of the p.R336C protein, which was absent upon pre-incubation with Cys. The WT and the p.R336H variant enzyme activity presented no increase with any of the tested compounds. Our results show that cysteamine and MEG are able to specifically improve the function of the CBS p.R336C variant, suggesting that any Arg-to-Cys substitution accessible to these small molecules may be converted back to a moiety resembling Arg.

ISOLATION AND MOLECULAR IDENTIFICATION OF POTENTIALLY PATHOGENIC Escherichia coli AND Campylobacter jejuni IN FERAL PIGEONS FROM AN URBAN AREA IN THE CITY OF LIMA, PERU / Isolamento e detecção de Escherichia coli e Campylobacter jejuni potencialmente patogênicos em pombos selvagens de uma área urbana na cidade de Lima, Perú

SUMMARY Feral pigeons (Columbia livia) live in close contact with humans and other animals. They can transmit potentially pathogenic and zoonotic agents. The objective of this study was to isolate and detect strains of diarrheagenic Escherichia coli and Campylobacter jejuniof urban feral pigeons from an area of Lima, Peru. Fresh dropping samples from urban parks were collected for microbiological isolation of E. coli strains in selective agar, and Campylobacterby filtration method. Molecular identification of diarrheagenic pathotypes of E.coliand Campylobacter jejuni was performed by PCR. Twenty-two parks were sampled and 16 colonies of Campylobacter spp. were isolated. The 100% of isolates were identified as Campylobacter jejuni. Furthermore, 102 colonies of E. coli were isolated and the 5.88% resulted as Enteropathogenic (EPEC) type and 0.98% as Shiga toxin-producing E. coli (STEC). The urban feral pigeons of Lima in Peru can act as a reservoir or carriers of zoonotic potentially pathogenic enteric agents.

RESUMO Os pombos selvagens (Columbia livia) vivem em estreito contato com os seres humanos e outros animais. Podem transmitir agentes potencialmente patogênicos e zoonóticos. Os objetivos deste estudo foram isolar e detectar cepas de Escherichia coli diarreiogênica e Campylobacter jejuni de pombos selvagens urbanos de uma área de Lima, Peru. Amostras de fezes frescas foram coletadas em parques urbanos para o isolamento microbiológico para cepas de E. coli em ágar seletivo e Campylobacterpor método de filtração. Identificação molecular de patótipos diarreiogênicos de E. coli e Campylobacter jejuni foi realizado por PCR. Vinte e dois parques foram amostrados e 16 colônias de Campylobacter spp. foram isolados. O 100% dos isolados foram identificados como Campylobacter jejuni. Além disso, 102 colônias de E. coli foram isoladas e 5,88% resultaram como tipo enteropatogênico (EPEC) e 0,98% como produtora de toxina Shiga (STEC). Os pombos selvagens urbanos de Lima no Peru podem atuar como reservatório ou ser portador de agentes zoonóticos entéricos potencialmente patogênicos.

Insights into the regulatory domain of cystathionine Beta-synthase: characterization of six variant proteins.

Cystathionine beta-synthase (CBS) catalyzes the formation of cystathionine from homocysteine and serine. CBS is allosterically activated by S-adenosylmethionine (SAM), which binds to its C-terminal regulatory domain. Mutations in this domain lead to variants with high residual activity but lacking SAM activation. We characterized six C-terminal CBS variants (p.P427L, p.D444N, p.V449G, p.S500L, p.K523Sfs*18, and p.L540Q). To understand the effect of C-terminal mutations on the functional/structural properties of CBS, we performed dynamic light scattering, differential scanning fluorimetry, limited proteolysis, enzymatic characterization, and determination of SAM-binding affinity. Kinetic data confirm that the enzymatic function of these variants is not impaired. Although lacking SAM activation, the p.P427L and p.S500L were able to bind SAM at a lower extent than the wild type (WT), confirming that SAM binding and activation can be two independent events. At the structural level, the C-terminal variants presented various effects, either showing catalytic core instability and increased susceptibility toward aggregation or presenting with similar or higher stability than the WT. Our study highlights as the common feature to the C-terminal variants an impaired binding of SAM and no increase in enzymatic activity with physiological concentrations of the activator, suggesting the loss of regulation by SAM as a potential pathogenic mechanism.

Reduced response of Cystathionine Beta-Synthase (CBS) to S-Adenosylmethionine (SAM): Identification and functional analysis of CBS gene mutations in Homocystinuria patients.

A reduced response of cystathionine beta-synthase (CBS) to its allosteric activator S-adenosylmethionine (SAM) has been reported to be a cause of CBS dysfunction in homocystinuria patients. In this work we performed a retrospective analysis of fibroblast data from 62 homocystinuria patients and found that 13 of them presented a disturbed SAM activation. Their genotypic background was identified and the corresponding CBS mutant proteins were produced in E. coli. Nine distinct mutations were detected in 22 independent alleles: the novel mutations p.K269del, p.P427L, p.S500L and p.L540Q; and the previously described mutations p.P49L, p.C165Rfs*2, p.I278T, p.R336H and p.D444N. Expression levels and residual enzyme activities, determined in the soluble fraction of E. coli lysates, strongly correlated with the localization of the affected amino acid residue. C-terminal mutations lead to activities in the range of the wild-type CBS and to oligomeric forms migrating faster than tetramers, suggesting an abnormal conformation that might be responsible for the lack of SAM activation. Mutations in the catalytic core were associated with low protein expression levels, decreased enzyme activities and a higher content of high molecular mass forms. Furthermore, the absence of SAM activation found in the patients' fibroblasts was confirmed for all but one of the characterized recombinant proteins (p.P49L). Our study experimentally supports a deficient regulation of CBS by SAM as a frequently found mechanism in CBS deficiency, which should be considered not only as a valuable diagnostic tool but also as a potential target for the development of new therapeutic approaches in classical homocystinuria.

Protein arginine methylation is more prone to inhibition by S-adenosylhomocysteine than DNA methylation in vascular endothelial cells.

Methyltransferases use S-adenosylmethionine (AdoMet) as methyl group donor, forming S-adenosylhomocysteine (AdoHcy) and methylated substrates, including DNA and proteins. AdoHcy inhibits most methyltransferases. Accumulation of intracellular AdoHcy secondary to Hcy elevation elicits global DNA hypomethylation. We aimed at determining the extent at which protein arginine methylation status is affected by accumulation of intracellular AdoHcy. AdoHcy accumulation in human umbilical vein endothelial cells was induced by inhibition of AdoHcy hydrolase by adenosine-2,3-dialdehyde (AdOx). As a measure of protein arginine methylation status, the levels of monomethylarginine (MMA) and asymmetric and symmetric dimethylated arginine residues (ADMA and SDMA, respectively) in cell protein hydrolysates were measured by HPLC. A 10% decrease was observed at a 2.5-fold increase of intracellular AdoHcy. Western blotting revealed that the translational levels of the main enzymes catalyzing protein arginine methylation, protein arginine methyl transferases (PRMTs) 1 and 5, were not affected by AdoHcy accumulation. Global DNA methylation status was evaluated by measuring 5-methylcytosine and total cytosine concentrations in DNA hydrolysates by LC-MS/MS. DNA methylation decreased by 10% only when intracellular AdoHcy concentration accumulated to 6-fold of its basal value. In conclusion, our results indicate that protein arginine methylation is more sensitive to AdoHcy accumulation than DNA methylation, pinpointing a possible new player in methylation-related pathology.

Asymmetric dimethylarginine in adults with cystathionine ß-synthase deficiency.

In hyperhomocysteinemia (HHcy), an independent risk factor for cardiovascular diseases, endothelial dysfunction due to reduced bioavailability of nitric oxide is a consistent finding. However, the underlying mechanisms remain unknown. Increased levels of the nitric oxide synthase inhibitor asymmetric dimethylarginine (ADMA) have been associated with HHcy, and may contribute, at least in part, for the homocysteine-induced endothelial dysfunction, but whether cystathionine ß-synthase (CBS) deficiency is associated with increased ADMA has hardly been investigated. To address this question, we measured total homocysteine (tHcy), ADMA and symmetric dimethylarginine (SDMA) in plasma of 22 adult CBS deficient patients, using established HPLC techniques. Results showed that in CBS deficient patients with elevated levels of tHcy (median (total range): 33 (14-237) µmol/L), both ADMA and SDMA levels were normal. Moreover, tHcy and ADMA concentrations were not correlated (r(s)=0.017, p=0.94). Our results favor the hypothesis that the negative vascular effects of HHcy have an ADMA-independent etiology.

Síndrome de vaciamiento rápido en pacientes sometidos a resección gástrica / Dumping syndrome in patients submitted to gastric resection

BACKGROUND: We undertook this study to establish the incidence of dumping syndrome after partial or total gastric resection and its association with patient's preoperative nutritional status as well as the clinical behavior with dietary management during a short-term follow-up period. METHODS: This was a prospective study of consecutive patients >30 years of age and who were submitted to gastrectomy for gastric cancer or complicated ulceropeptic disease during a 48-month period in a highly specialized hospital. RESULTS: A total of 42 patients were evaluated with a slight female predominance (n = 22, 52.4%). Twenty-nine cases (69%) had subtotal gastrectomy and 13 (31%) had a total gastrectomy. Patients had a medium age of 54.38 +/- 7.56 vs. 66 +/- 13.99 years, respectively (p = 0.034). Reconstruction techniques were Roux-en-Y gastrojejunostomy in 70% and Roux-en-Y esophagojejunostomy in 28.5%. We found dumping syndrome in 45% of the cases associated with acute or chronic undernutrition (p = 0.003). Fifty-three percent of the patients with dumping syndrome improved with adequate dietetic manipulation during a follow-up period of 211 days. CONCLUSIONS: Although the majority of reconstructions were performed with dysfunctionalized small bowel segments, the incidence of dumping syndrome was 45%. Patient's preoperative nutritional status influenced the presence of clinical manifestations. Adequate dietary management reduced, in 53% of the patients, the presence of dumping symptoms during a short-term follow-up period.

[Dumping syndrome in patients submitted to gastric resection]. / Síndrome de vaciamiento rápido en pacientes sometidos a resección gástrica.

BACKGROUND: We undertook this study to establish the incidence of dumping syndrome after partial or total gastric resection and its association with patient's preoperative nutritional status as well as the clinical behavior with dietary management during a short-term follow-up period. METHODS: This was a prospective study of consecutive patients >30 years of age and who were submitted to gastrectomy for gastric cancer or complicated ulceropeptic disease during a 48-month period in a highly specialized hospital. RESULTS: A total of 42 patients were evaluated with a slight female predominance (n = 22, 52.4%). Twenty-nine cases (69%) had subtotal gastrectomy and 13 (31%) had a total gastrectomy. Patients had a medium age of 54.38 +/- 7.56 vs. 66 +/- 13.99 years, respectively (p = 0.034). Reconstruction techniques were Roux-en-Y gastrojejunostomy in 70% and Roux-en-Y esophagojejunostomy in 28.5%. We found dumping syndrome in 45% of the cases associated with acute or chronic undernutrition (p = 0.003). Fifty-three percent of the patients with dumping syndrome improved with adequate dietetic manipulation during a follow-up period of 211 days. CONCLUSIONS: Although the majority of reconstructions were performed with dysfunctionalized small bowel segments, the incidence of dumping syndrome was 45%. Patient's preoperative nutritional status influenced the presence of clinical manifestations. Adequate dietary management reduced, in 53% of the patients, the presence of dumping symptoms during a short-term follow-up period.

Intracellular S-adenosylhomocysteine increased levels are associated with DNA hypomethylation in HUVEC.

Hyperhomocysteinemia is a risk factor for atherosclerosis and vascular disease; however, the mechanism underlying this association remains poorly understood. Increased levels of intracellular S-adenosylhomocysteine (AdoHcy), secondary to homocysteine-mediated reversal of the AdoHcy hydrolase reaction, have been associated with reduced DNA methylation patterns and pointed as responsible for the hyperhomocysteinemia-related endothelial dysfunction. Methylation is an epigenetic feature of genomic DNA, which leads to alterations in gene expression. So far, the effect of intracellular AdoHcy accumulation on DNA methylation patterns has not yet been fully substantiated by experimental evidence. The present study was designed to evaluate, in cultured endothelial cells, the effect of AdoHcy accumulation on genomic global DNA methylation status. Experimental intracellular accumulation of AdoHcy was induced by adenosine-2,3-dialdehyde (ADA), an inhibitor of AdoHcy hydrolase. Increased concentrations of inhibitor were tested, and unsupplemented medium incubations were used as controls. Cytosolic and nuclear fractions were obtained from trypsinized cells after 72 h of incubation. Total homocysteine concentration was quantified (culture medium and cytosolic fractions) by high-performance liquid chromatography (HPLC). S-Adenosylmethionine and AdoHcy concentrations were measured (cytosolic fractions) by stable-isotope dilution LC-tandem mass spectrometry method. Genomic DNA was obtained from the nuclear fraction, and global DNA methylation status was evaluated by the cytosine extension assay. The results showed that supplementation of the culture medium with ADA had no cytotoxic effect and increased the intracellular AdoHcy concentration in a dose-dependent manner. A significant negative correlation was observed between intracellular AdoHcy and genomic DNA methylation status. These findings strongly point to the importance of AdoHcy as a pivotal biomarker of genomic DNA methylation status.

Increased homocysteine and S-adenosylhomocysteine concentrations and DNA hypomethylation in vascular disease.

BACKGROUND: The pathogenic mechanism of homocysteine's effect on cardiovascular risk is poorly understood. Recent studies show that DNA hypomethylation induced by increases in S-adenosylhomocysteine (AdoHcy), an intermediate of Hcy metabolism and a potent inhibitor of methyltransferases, may be involved in homocysteine-related pathology. METHODS: We measured fasting plasma total Hcy (tHcy), AdoHcy, and S-adenosylmethionine (AdoMet) and methylation in leukocytes in 17 patients with vascular disease and in 15 healthy, age- and sex-matched controls. RESULTS: Patient with vascular disease had significantly higher plasma tHcy and AdoHcy concentrations and significantly lower plasma AdoMet/AdoHcy ratios and genomic DNA methylation. AdoMet concentrations were not significantly different between the two groups. More than 50% of the patients fell into the highest quartiles of plasma tHcy, AdoHcy, and [(3)H]dCTP incorporation/ micro g of DNA (meaning the lowest quartile of DNA methylation status) and into the lowest quartile of the AdoMet/AdoHcy ratios of the control group. Plasma tHcy was significantly correlated with plasma AdoHcy and AdoMet/AdoHcy ratios (n = 32; P < 0.001). DNA methylation status was significantly correlated with plasma tHcy and AdoHcy (n = 32; P < 0.01) but not with plasma AdoMet/AdoHcy ratios. CONCLUSION: Global DNA methylation may be altered in vascular disease, with a concomitant increase in plasma tHcy and AdoHcy.