Prediction of the antigenic regions in eight RhD variants identified by computational biology.

BACKGROUND AND OBJECTIVES: Changes in RHD generate variations in protein structure that lead to antigenic variants. The classical model divides them into quantitative (weak and Del) and qualitative (partial D). There are two types of protein antigens: linear and conformational. Computational biology analyses the theoretical assembly of tertiary protein structures and allows us to identify the 'topological' differences between isoforms. Our aim was to determine the theoretical antigenic differences between weak RhD variants compared with normal RhD based on structural analysis using bioinformatic techniques. MATERIALS AND METHODS: We analysed the variations in secondary structures and hydrophobicity of RHD*01, RHD*01W.1, W2, W3, RHD*09.03.01, RHD*09.04, RHD*11, RHD*15 and RHD*21. We then modelled the tertiary structure and calculated their probable antigenic regions, intra-protein interactions, displacement and membrane width and compared them with Rhce. RESULTS: The 10 proteins are similar in their secondary structure and hydrophobicity, with the main differences observed in the exofacial coils. We identified six potential antigenic regions: one that is unique to RhD (R3), one that is common to all D (R6), three that are highly variable among RhD isoforms (R1, R2 and R4), one that they share with Rhce (R5) and two that are unique to Rhce (Ra and Rbc). CONCLUSION: The alloimmunization capacity of these subjects could be explained by the variability of the antigen pattern, which is not necessarily recognized or recognized with lower intensity by the commercially available antibodies, and not because they have a lower protein concentration in the membrane.

Use of computational biology to compare the theoretical tertiary structures of the most common forms of RhCE and RhD.

BACKGROUND AND OBJECTIVES: Computational biology analyses the theoretical tertiary structure of proteins and identifies the 'topological' differences between RhD and RhCE. Our aim was to identify the theoretical structural differences between the four isoforms of RhCE and RhD using computational biological tools. MATERIALS AND METHODS: Physicochemical profile was determined by hydrophobicity and electrostatic potential analysis. Secondary and tertiary structures were generated using computational biology tools. The structures were evaluated and validated using Ramachandran algorithm, which calculates the single score, p-value and root mean square deviation (RMSD). Structures were overlaid on local refinement of 'RhAG-RhCE-ANK' (PBDID 7uzq) and RhAG to compare their spatial distribution within the membrane. RESULTS: All proteins differed in surface area and electrostatic distance due to variations in hydrophobicity and electrostatic potential. The RMSD between RhD and RhCE was 0.46 ± 0.04 Å, and the comparison within RhCE was 0.57 ± 0.08 Å. The percentage of amino acids in the hydrophobic thickness was 50.24% for RhD while for RhCE it ranged between 73.08% and 76.68%. The RHAG hydrophobic thickness was 34.2 Å, and RhCE's hydrophobic thickness was 33.83 Å. We suggest that the C/c antigens differ exofacially at loops L1 and L2. For the E/e antigens, the difference lies in L6. By contrast, L4 is the same for all proteins except Rhce. CONCLUSION: The physicochemical properties of Rh proteins made them different, although their genes are homologous. Using computational biology, we model structures with sufficient precision, similar to those obtained experimentally. An amino acid variation alters the folding of the tertiary structure and the interactions with other proteins, modifying the electrostatic environment, the spatial conformations and therefore the antigenic recognition.

Analysis of a Novel Mexican Variant of the HBB Gene Associated with ß-Thalassemia Using Bioinformatic Tools.

We present a study performed on 54 unrelated subjects, with and without thalassemic features. Two primer pairs were proposed to perform Sanger sequencing of the complete HBB gene. The bioinformatic analysis was performed taking advantage of the availability of free online tools. In the sample, we found 11 variants, 10 reported, and one novel. Among the variants found, six are clinically important: three encode a premature stop codon [codon 39 (C>T) (HBB: c.118C>T); IVS-II-1 (G>A) (HBB: c.315+1G>A), and one not reported], a double substitution within the same allele [Hb Borås (HBB: c.266T>G) and Hb Santa Giusta Sardegna (HBB: c.282T>C)], and one whose pathogenicity is not yet defined [Hb Fannin-Lubbock I (HBB: c.359G>A)]. Even though the variants Hb Borås and Hb Santa Giusta Sardegna have been described, there is no report of their combined occurrence on the same allele, which could cause hemolytic anemia. Although the p.Leu88Arg and p.Cys93Trp variants do not alter the final length of the protein, the bioinformatic results suggest that there are differences in the tertiary structure of ß-globin genes, mainly affecting helices E and F, being the motifs of interaction with the heme group. The novel variant is a 4 bp insertion that modifies the open reading frame, changing the last amino acid residue and causing a premature stop codon (HBB: c.291-294insGCAC). The variant was associated with ß-thalassemia (ß-thal). Bioinformatic analysis made it possible to predict the consequences that the new variant of the HBB gene caused on the ß-globin tertiary structure.

Parámetros de laboratorio de importancia en el manejo de pacientes con COVID-19 / Laboratory parameters of importance in the management of COVID-19 patients

Resumen El coronavirus 2 del síndrome respiratorio agudo grave (SARS-CoV-2) detona el padecimiento la enfermedad por coronavirus 2019 (COVID-19), poniendo en riesgo de muerte a la población vulnerable. El laboratorio clínico enfrenta un reto para el diagnóstico, seguimiento, pronóstico y evaluación de los tratamientos, que se ofrecen a los enfermos de COVID-19. Nuestro objetivo es mostrar al lector un resumen de los principales cambios en los parámetros que se estudian en los laboratorios clínicos. Material y métodos: Se hizo una búsqueda bibliográfica cruzando los términos COVID-19 y laboratorio clínico. Se analizaron las publicaciones relevantes por los miembros del Comité de Trombosis y Hemostasia-AMEH A.C. y se plasmaron las pruebas que a criterio de los participantes destacan por la relación entre la información que proporcionan respecto al seguimiento, pronóstico y evaluación al tratamiento. Resultados: Se recomienda solicitar una citometría hemática (recuento de células blancas, relación neutrófilo:linfocito), química sanguínea (transaminasas, bilirrubinas, albúmina, urea, creatinina, glucosa, lactato deshidrogenasa), pruebas de coagulación (tiempo de protrombina, tiempo de tromboplastina parcial activado, fibrinógeno y dímeros D) y pruebas especiales (proteína C reactiva, ferritina, procalcitonina, troponina).

Abstract The severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) triggers the coronavirus disease 2019 (COVID-19), putting the vulnerable population at risk of death. The clinical laboratory faces a challenge for the diagnosis, monitoring, prognosis and evaluation of therapy with low molecular weight heparin. Our objective in this article is to offer a brief discuss of the main changes in the clinical parameters, studied on behalf of COVID-19 patients by a clinical laboratory. Material and methods: A bibliographic search was made crossing the terms COVID-19 and clinical laboratory. Relevant publications were analyzed by the members of the Committee for Thrombosis and Hemostasis-AMEH A.C. The relevant articles were discussed, and the clinical tests discussed in the article are those, that meet the criteria of providing information referring to the follow-up, prognosis and evaluation of treatment against the lower cost. Results: It is recommended to request a blood count (white cell count, neutrophyl/ lymphocytes ratio), clinical chemistry (transaminases, bilirubin, albumin, urea, creatinine, glucose, lactate dehydrogenase), coagulation tests (prothrombin time, activated partial thromboplastin time, fibrinogen, DD dimers) and special tests (C-reactive protein, ferritin, procalcitonin, troponins).

[Molecular identification of Hemoglobin D Punjab in cases detected in two families]. / Identificación molecular de la hemoglobina D Punjab en dos familias.

BACKGROUND: Hemoglobin D Punjab is the world most common variant hemoglobin D; in Mexico there are reports of isolated cases. Our goal is to present the clinical and molecular study in two families with HbD Punjab. The objective was to submit molecular diagnosis of two families with Hb D Punjab and clinical features. CLINICAL CASE: Family 1: neonate with maternal history of HbS carrier. Father and sister with natural variants for the evaluated mutations, mother, brother and index case were heterozygous for HbD Punjab. Family 2: neonate with positive neonatal screening for detection of abnormal hemoglobins. Mother and index case were heterozygous for HbD Punjab, homozygous for HFE H63D, and Gilbert's syndrome UGT1A1*28 heterozygous. Father heterozygous for HFE H63D and sister homozygous for such mutation. The study of two families for HbD Punjab, HbS, ß-thalassemia, HFE and Gilbert syndrome was performed by real time PCR. CONCLUSION: The molecular identification of HbD Punjab is an accessible methodological proposal that can adequately distinguish carriers subjects; through this method two additional cases, one initially identified as HbS.

Introducción: la hemoglobina D Punjab (HbD Punjab) es la variante mundial más frecuente de hemoglobina D. En México se tienen reportes de casos aislados. El objetivo es presentar el diagnóstico molecular de dos familias con HbD Punjab y sus características clínicas. Casos clínicos: familia 1: neonato cuya madre era portadora de HbS. El padre y la hermana tenían variante natural para las mutaciones evaluadas, madre, hermano y caso índice heterocigotos para HbD Punjab. Familia 2: neonato con tamiz neonatal positivo para la detección de hemoglobinas anormales. Madre y caso índice fueron heterocigotos para HbD Punjab, homocigotos para HFE H63, y heterocigotos para síndrome de Gilbert UGT1A1*28. Padre, heterocigoto para HFE H63D, y hermana homocigoto para dicha mutación. El estudio de las dos familias para HbD Punjab, HbS, beta-talasemia, HFE y síndrome de Gilbert se hizo mediante PCR en tiempo real. Conclusión: la identificación molecular de la HbD Punjab es una propuesta metodológica accesible y permite diferenciar adecuadamente a los portadores. A través de esta metodología se identificaron dos casos adicionales en nuestro país, uno de ellos identificado inicialmente como HbS.

[Molecular identification of glucose-6-phosphate dehydrogenase (G6PD) detected in neonatal screening]. / Identificación molecular de la glucosa-6-fosfato deshidrogenasa (G6PD) detectada en el tamiz neonatal.

AIMS: To present the strategy of identifying the molecular variants of G6PD detected in neonatal screening (NS). MATERIAL AND METHODS: We present a series of incident cases of newborns positive for G6PD deficiency detected in NS. From nuclear DNA with the methodology of real-time PCR we sought molecular G6PD variants: G202A, A376G, T968C and C563T. RESULTS: Of a total of 21,619 neonates, 41 cases were reactive in NS for G6PD (189.6/100,000 RN screened rate), 34 cases confirmed the molecular variant of G6PD (157.3/100,000 RN screened rate). The most frequent allele combination G202A/A376G (G6PD ratio and median activity, 0.460 and 1.72 ± 0.35 U/g Hb, respectively), followed by G202A (0.170 and 1.74 ± 0.27 U/g Hb) and A376G/T968C (ratio 0.150 and 1.10 ± 0.44 U/g Hb). The T968C allelic variant showed lower enzyme activity than the rest (1.1 ± 0.4; p = 0.02). Two women were detected with G6PD deficiency with G202A/A376G and G202A variant. CONCLUSIONS: African alleles were prevalently detected in neonatal screening. This strategy allows the identification of molecular variants involved in 80% of cases.

Variaciones en el recuento diferencial leucocitario desde el nacimiento hasta los primeros dos meses de edad en nacidos a término con bajo riesgo perinatal / Variations in the differential leukocyte count from birth through the first two months of age in term infants with low perinatal risk

Introducción: La presencia de neutropenia o neutrofilia se observa en la evaluación del neonato con sospecha de sepsis; sin embargo, en ausencia de infección sistémica, su incidencia no ha sido estimada en nuestra población. Material y métodos: En una cohorte de recién nacidos a término de bajo riesgo perinatal, seguidos desde el nacimiento hasta los dos meses de edad; se evaluaron la cuenta de leucocitos, neutrófilos (NT), linfocitos totales (LT) y la frecuencia de neutropenia a estas edades. Resultados: Se incluyeron 110 recién nacidos de bajo riesgo perinatal, evaluados al nacer, al mes y a los dos meses de edad. Los leucocitos totales promedio fueron de 18,950/µL (IC 95% 10,255 a 29,170), 11,250/µL (6,030-17,045) y 9,750/µL (6,055-17,900); los linfocitos totales de 7,442/µL (4,736-11,326), 8,198/µL (2,625-12,559) y 7,478/µL (4,276-12,782); los neutrófilos totales de 7,818/µL (2,853-13,809), 2,112 (927-7,021) y 1,944 (737-3,618), respectivamente. La incidencia de neutropenia significativa (< 750/L) fue del 0.9 al nacimiento, 2.7 al mes y 5.5% a los dos meses de edad. La neutrofilia (> 9,500/L) se presentó en el 35.5% de los neonatos al nacimiento, pero en ningún caso hacia el resto de las edades de estudio. Conclusiones: La presencia de neutropenia o neutrofilia es un cambio fisiológico en el neonato o lactante menor de bajo riesgo. Este hecho debe ser considerando en su interpretación particular en la evaluación de la sepsis a estas edades.

Introduction: The presence of neutropenia or neutrophilia is observed in the evaluation of the neonate with suspected sepsis; however, its incidence has not been estimated in our population in the absence of systemic infection. Material and methods: A cohort of newborns at term, with low perinatal risk, were followed from birth to two months of age, leukocyte count, neutrophils (NT), and total lymphocyte (TL) were evaluated; the frequency of neutropenia at these ages was determined. Results: We included 110 infants; the mean values of the total leukocyte count were 18,950/µL (CI 95% 10,255-29,170); 11,250/µL (6,030-17,045) and 9,750/µL (6,055-17,900); total lymphocytes, 7,442/µL (4,736 to 11,326), 8,198/µL (2,625-12,559) and 7,478/µL (4,276-12,782); total neutrophil count, 7,818/µL (2,853 to 13,809), 2,112 (927-7,021) and 1,944 (737-3,618) at birth, one and two months, respectively. Significant neutropenia (< 750/L) was 0.9 at birth, 2.7 at one month and 5.5% at two months of age. Neutrophilia (> 9,500/L) was present in 35.5% of infants at birth, but in none in the first and second months of age. Conclusions: The presence of neutropenia or neutrophilia is a physiological change in low-risk infants. This fact must be considered when evaluating sepsis at this age.

Evaluación del estado en la reserva neonatal de hierro y las mutaciones del gen HFE / Assessment of neonatal iron store and HFE gene mutations

Introducción: Se presenta la evaluación de la asociación entre la reserva de hierro (Fe) y los polimorfismos del gen de la hemocromatosis (HFE) en neonatos de alto riesgo perinatal. Métodos: Se incluyó una serie de neonatos de alto riesgo perinatal en los que se evaluó la reserva de Fe con la medición de la ferritina sérica (FS). Se dividieron en tres grupos: sobrecarga de Fe (SoFe), con FS >1,000 µg/l; reserva normal de Fe, con FS de 154-1,000 µg/l; y reserva baja de Fe, con FS <154 µg/l. Mediante PCR en tiempo real se buscaron las mutaciones C282Y, H63D y S65C del gen HFE. Resultados: Se estudiaron 97 neonatos. De ellos, 24 casos presentaron SoFe (proporción 0.247) y FS de 1,789 µg/l (IC 95% 1,376-2,201); 36 casos, reserva normal de FS (0.371), FS de 461 µg/l (389-533); y 37 casos, reserva baja de FS (0.381) y FS 82 µg/l (69-96). No hubo casos detectados para las mutaciones C282Y o S65C. Se identificó la variante H63D HFE en 18 neonatos (frecuencia génica de 0.185): la condición de heterocigoto (H63D/WT) en doce casos (frecuencia génica 0.124) y de homocigoto (H63D/H63D) en seis casos (frecuencia génica 0.062). La frecuencia alélica de H63D fue de 0.092. Los variante H63D HFE no mostró asociación con los neonatos de reserva normal de Fe contra reserva baja (OR 1.2; IC 95% 0.3-4.3) ni los de reserva normal contra neonatos con SoFe (OR 2.5; 0.7-9.2). Conclusiones: Cerca del 25% de neonatos de alto riesgo tendrá sobrecarga de Fe. Aún con el posible sesgo de selección, las variantes del gen HFE no influyen sobre el estado de la reserva de Fe.

Background: The association between iron stores (Fe) and HFE gene polymorphisms on high-risk neonates is shown. Methods: We included newborns with high perinatal risk. Newborns were divided into three groups for measurements of serum ferritin (SF): iron overload (IO) with SF 1000 µg/L, normal iron stores (NIS) with SF 154-1000 µg/L and low iron stores (LIS) with SF <154 µg/L. We used real-time PCR for identification of polymorphisms C282Y, H63DE, and S65C of the HFE gene. Results: We studied 97 newborns with IO in 24 cases (ratio 0.247) and SF 1789 µg/L (95% CI 1376-2201), NIS in 36 cases (0.371), and SF of 461 µg/L (389-533) and LIS in 37 cases (0.381) and SF 82 µg/L (69-96). There were no cases detected for C282Y or S65C mutations. We identified 18 neonates with H63D HFE variant (gene frequency 0.185) with heterozygous condition (H63D/ WT) in 12 cases (gene frequency 0.124) and homozygote (H63D/H63D) in six cases (gene frequency 0.062). H63D allele frequency was 0.092. The HFE H63D variant showed no association for comparing infants with NIS vs. LIS (OR 1.2, 95% CI 0.3-4.3) and NIS vs. IO newborn infant (OR 2.5, 0.7-9.2). Conclusions: In high-risk neonates ∼25% show IO even with the possible selection bias. HFE gene variants do not influence on the neonatal iron stores.

Association between genotype and plasma levels of thrombin-activated fibrinolysis inhibitor (TAFI) in the development of preeclampsia.

INTRODUCTION: The objective was to evaluate if thrombin-activated fibrinolysis inhibitor (TAFI) polymorphisms (G505A, C1040T, and G-438A), and TAFIa plasma levels are associated with preeclampsia. MATERIALS AND METHODS: In a case-control study design, we evaluated preeclampsia patients and women with uncomplicated pregnancies. The TAFI polymorphisms were determined by real-time PCR method, and TAFIa plasma levels were established with a chromogenic assay. RESULTS: We included 87 women in each group. The TAFIa levels in the preeclampsia group were 20.4 µg/mL (CI 95% 17.3-23.5), while in the control group, they were significantly lower: 13.3 µg/mL (12.0-14.5, p 0.003). There were no differences in the genotype distribution or allelic frequency of TAFI polymorphisms between the two groups. In preeclampsia patients and controls heterozygous for the G505A polymorphism, the TAFIa values were 22.8 (16.7-28.9 µg/mL) and 13.2 (11.3-15.0 µg/mL, p 0.019), respectively. In G505A homozygous polymorphism the TAFIa values were 25.7 (18.7-32.6 µg/mL) and 13.5 (1.6-21.9 µg/mL, p 0.041), respectively. In the C1040T and G-438A TAFI wild type polymorphisms, the TAFIa values were 18.3 (12.5-23.9 µg/mL) and 11.5 (9.9-35.0, p 0.033), and 19.4 (10.9-27.9 µg/mL) and 12.5 (10.8-14.2 µg/mL, p 0.006), respectively, without differences in other genotypes. CONCLUSIONS: Preeclampsia by itself may be responsible for the increase in TAFIa values rather than the presence of polymorphisms.

[RH hybrid box identification in subjects with Rh negative phenotype from Mexico's Valley]. / Identificación de la caja RH híbrida en el fenotipo Rh negativo de sujetos del valle de México.

BACKGROUND: The prevalence of the RhD and RhCE gene alleles is related to the ethnic mixture. The aim of this report is to describe the predominant molecular mechanisms in RhD negative subjects residents from Mexico's valley according to the phenotype of RhCE. METHODS: Blood samples from RhD negative women and men were studied. The RhD/RhCE phenotype was identified by hemagglutination and Rh hybrid box by PCR-FRLP with PstI. RESULTS: 216 subjects were included. The RhD phenotypes were ccdee in 179 cases (82.8%), Ccdee in 15 cases (11.6%), ccdEe in seven (3.2%), CcdEe in four (1.9%), and CcdEE in a single subject (0.5%). In five cases, RhD hybrid box was not amplified (2.3%), 21 cases were hemizygotes (9.7%), and 188 cases homozygotes (87%), for RhD hybrid box. The homozygote condition was more frequent in those individuals with phenotype ccdee (87%). The allelic frequency of RhD hybrid box was 0.928. The frequency of Rhcc haplotype was higher in those subjects homozygotes for RhD hybrid box (chi2 = 4.658, p < 0.05). CONCLUSIONS: In this population, RhD gene deletion is the main molecular mechanism to generate to RhD negative condition and this depends on the European mixture.

Utilidad de la prueba directa de Coombs en el tamiz neonatal / Usefulness of direct antiglobulin test in neonatal screening

Introducción. La ictericia es un evento clínico frecuente que se presenta en los recién nacidos; las causas más frecuentes involucradas en la enfermedad hemolítica del recién nacido (EHRN) continúan siendo la incompatibilidad al sistema ABO y la isoinmunización a RhD. La prueba directa de Coombs (PDC) permite identificar la presencia de anticuerpos antieritrocitarios del isotipo IgG, provenientes del suero materno en la superficie de los eritrocitos del feto o neonato. Objetivo: presentar los resultados y especificidad de la prueba directa de Coombs (PDC) como prueba en el tamiz neonatal. Métodos. Consecutivamente se incluyeron a recién nacidos no seleccionados. En las primeras horas de vida se determinó el grupo sanguíneo ABO/RhD y la PDC con suero poliespecífico y monoespecíficos (anti-IgG y C3b/C3d). Resultados. Se incluyeron 5 007 recién nacidos, la PDC positiva se documentó en 181 neonatos (3.6%). Los casos del grupo sanguíneo A, B o AB, mostraron mayor riesgo de tener PDC positiva que los del grupo O (razón de momios 2.3, intervalo de confianza 95% 1.7-3.1). La PDC positiva se presentó en 3.5% de los neonatos RhD positivo y en 1.9% de los RhD negativo; 72.9% de los neonatos con PDC positiva tuvieron titulaciones de 1:2, 1 ;4 y 1:8, con el 33.1, 19.9 y 19.9%, respectivamente. En los neonatos con PDC positiva se pudo establecer el isotipo anti-IgG en 117 casos (64.6%), complemento sólo en 6 casos (3.3%), la combinación de ambas en 6 casos (3.3%) y en 52 neonatos (28.8%), con PDC poliespecífico positivo, no se pudo identificar la especificidad de la reacción. Conclusión. La prevalencia elevada (3.6 %) de PDC positiva en la detección neonatal rutinaria, apoya la indicación de incorporar dicha prueba al tamiz neonatal, independientemente del grupo ABO y Rh materno. Queda por establecer su beneficio en la detección temprana de la ictericia neonatal.

Background. Neonatal jaundice is a clinical event frequently present in newborns. The causes most frequently involved in hemolytic disease of newborn (HDN) are still the incompatibilities to the ABO/Rh blood system. Direct Coombs test (or direct antiglobulin test, DAT) allows identification of the presence of red blood cell antibodies (IgG isotype) coming from the maternal serum on the surface of the fetus erythrocytes. The purpose of this study is to show the results and specificity of DAT as screening in newborn infants. Methods. We studied unselected neonates in a cross-sectional design. During the early hours of life, we determined ABO/Rh and DAT with poly- and monospecific reagents (anti-IgG and C3b/C3d). Results. We included 5 007 newborns; 181 cases (3.6%) were DAT positive. Newborns with A, B or AB blood groups showed an increased association of being DAT positive than group O (OR 2.3, 95% Cl 1.7-3.1). DAT was positive in 3.5% of RhD-positive infants and 1.9% of RhD-negative infants. In six DAT-positive cases, 117 cases (64.6%) had anti-IgG bound to red cell membrane, complement in six cases (3.3%), and 52 newborns (28.8%) were polyspecific DAT positive and monospecific DAT negative. Conclusions. The high prevalence (3.6%) of DAT-positive cases in routine neonatal detection supports the indication to incorporate DAT into neonatal screening, regardless of the mother's blood group. The benefit of early intervention in neonatal jaundice remains to be established.

Association of HFE mutations (C282Y and H63D) with iron overload in blood donors from Mexico City.

BACKGROUND AND OBJECTIVE: Iron overload has been associated with HFE mutations (C282Y and H63D). We investigated the association between these mutations and high serum ferritin in a sample of healthy adult men. DESIGN AND METHODS: We enrolled unrelated blood donors from three hospitals in Mexico City in a crosssectional study. Serum ferritin (SF) was determined to define iron overload, and HFE gene mutations were identified by PCR-RFLP. RESULTS: We evaluated 2524 male blood donors and included 246 individuals for each group. We identified 108 individuals with HFE gene mutation, 20.5 % were heterozygote (wt/H63D or wt/C282Y) and the remaining homozygote (H63D/ H63D). The genotype wt/C282Y was observed in two cases, none cases with C282Y/C282Y. The allelic frequency of H63D and C282Y was 0.115 and 0.002, respectively. We observed different association for H63D allele with iron overload (OR 1.54, CI 95 %1.16-2.03) and none in allele C282Y. Although values averages were different, the extreme dispersion of serum ferritin not showed statistically significant differences between H63D and C282Y alleles and ferritin concentrations. CONCLUSIONS: The male unrelated blood donors from Mexico City with iron overload prevalence of 13.8% hold similarities with other populations from Europe o America continent, respecting the allele frequency H63D. Nevertheless, allele frequency C282Y is lower than that observed in descendents from northern Europe. We have not observed statistic difference of SF or iron overload frequency by effect of both alleles.

Changes of coagulation inhibitors and fibrinolysis system in newborn infants with transitory neonatal cholestasis.

INTRODUCTION: The hemostatic system in newborn is a dynamic evolving process health-status dependent. OBJECTIVE: To explore the changes in coagulation inhibitors and fibrinolytic system in newborn with neonatal cholestasis, according liver damage intensity. METHODS: In a cross-sectional design, we studied fibrinolysis and coagulation inhibitor proteins, and serum ferritin (SF) in patients with neonatal cholestasis. We stratified the cases according the results of ALT or AST < 100 UI (group I) and > or =100 U/L (group II). RESULTS: We included 24 newborn, 8 for Group I and 16 cases for group II. We documented statistical differences in ATIII values (43.4 vs 27.4%, p < 0,01), plasmin inhibitor (93.5 vs 63.6%, p < 0.01), SF (649 vs 1410 ug/L, p 0.01). The group II cases showed association with SF values (f 20.6, p < 0.01) and plasmin inhibitor (f 40.4, p < 0.01). AST and ALT were related significantly to ATIII concentrations and SF. DISCUSSION: We documented tendency to prothrombotic state (lower ATIII and greater plasmin inhibitor activity), with low plasminogen related to the intensity of liver dysfunction in neonatal cholestasis. We need to determine the role of iron overload in physiopathology of the disease.

Influencia de la velocidad de crecimiento y tipo de alimentación láctea sobre la reserva corporal de hierro a los dos meses de edad

Introducción. El objetivo del estudio es establecer la influencia que tiene la alimentación láctea y la velocidad de crecimiento sobre la reserva de hierro (Fe) a los 2 meses de edad. Material y métodos. En un estudio longitudinal y aleatorio, se incluyeron a neonatos de término sanos, registrando las variables de crecimiento (peso, longitud supina, perímetro cefálico), así como los valores de hemoglobina (Hb) y ferritina sérica (FS) al nacimiento, 1 y 2 meses de edad, de acuerdo al tipo de alimentación láctea (materna, sucedáneos o mixta). Resultados. Se incluyeron 127 casos, con 57, 14 y 56 lactantes para cada grupo de alimentación. No ocurrieron diferencias estadísticas en la velocidad de crecimiento, entre los 3 grupos. Los valores promedio de Hb fueron de 19.6, 18.8 y 19.1 g/dL, de 14.3, 13.3 y 13.7 g/dL y de 12.0, 11.7 y 11.7 g/dL, para la FS los valores promedio fueron de 333, 297 y 347 µg/L, de 253, 277 y 258 µg/L y al segundo mes de edad 237, 227 y 243 µg/L, sin diferencias estadísticas. La prevalencia global de deficiencia de Fe fue de 3.9%. Conclusiones. Los preparados comerciales de la leche, fortificados con 12 mg/L de Fe, no presentan ventaja adicional a la leche materna, la velocidad de crecimiento corporal, valores de Hb y reserva de Fe a los 2 meses de edad.

Introduction. The objective of this study was to establish the influence of milk feeding and growth velocity on iron store at 2 months of age. Material and methods. In a longitudinal and randomized study, we included to healthy newborn, anthropometric data (weight, height, and cephalic perimeter), hemoglobin (Hb), and serum ferritin (SF) values at birth, 1 and 2 months of age were obtain according to type of milk feeding received (breast, formula fortified with iron at 12 mg/L or mixed). Results. We included 127 cases, with 57, 14 and 56 newborn for each group. No statistical differences were observed in growth velocity. The values average of Hb were of 19.6, 18.8 and 19.1 g/dL, of 14.3, 13.3 and 13.7 g/dL and of 12.0, 11.7 and 11.7 g/dL, for the FS the values average were of 333, 297 and 347 µg/L, of 253, 277 and 258 µg/L and 2 month of age 237, 227 and 243 µg/L, without statistical differences. The global prevalence of iron deficiency was of 3.9%. Conclusions. Milk formulas, fortified with iron at 12 mg/L, do no represent additional advantage in body growth, Hb values and iron store.

Aislamiento e identificación de las células fetales presentes al nacimiento en la circulación materna / Identification at delivery of fetal cells in the maternal circulation

Introducción. El objetivo es presentar un modelo experimental para el aislamiento de las células fetales presentes en la circulación materna.Material y métodos. Se estudiaron a mujeres Rh negativo al final del evento obstétrico, recolectándose sangre venosa materna y del cordón umbilical del recién nacido. Se aislaron las células fetales en circulación materna mediante centrifugación con gradiente de separación, seguido de separación selectiva con perlas magnéticas marcadas anti-CD14 y anti-CD45, para posteriormente retirar los eritroblastos fetales con anti-CD71. A las células obtenidas en este último paso se les extrajo el ADN para amplificar mediante PCR, las regiones de los exones 4 y 5 de los genes RhD y RhCE. El fenotipo del Rh materno y neonatal se estableció mediante inmunohemaglutinación.Resultados. Se estudiaron 7 mujeres Rh negativo y sus respectivos hijos con fenotipo Rh positivo. De las células fetales aisladas se amplificaron en PCR dos bandas en electroforesis, una de 1 200 pares de bases (pb) para el gen RhD y otra de 600 pb para el gen RhCE. Los recién nacidos mostraron ambas bandas, mientras que las madres sólo presentaron la banda de 600 bp (Gen RhCE). Se obtuvo concordancia entre el fenotipo y el genotipo del RhD en 6 de 7 casos. En un caso no se obtuvo suficiente material celular para obtener ADN. Conclusión. Se muestra una estrategia que puede permitir el aislar células fetales de la circulación materna, abriendo una posibilidad de que posteriormente sea validado para el estudio del ADN mediante procedimientos sin riesgo fetal alguno.

Prevención de la isoinmunización materna al RhD, con gamma-globulina anti-D / Prevention of maternal RhD isoimmunization with anti-D gamma globulin

Objetivo. Presentar la experiencia institucional en la prevención de la isoinmunización al RhD, mediante el empleo de 150 µg de g-globulina anti-D en las mujeres Rh negativo. Material y métodos. Se registraron los antecedentes inmunohematológicos de los casos consecutivos de todas las mujeres Rh negativo que acudieron, para su atención médica, al Instituto Nacional de Perinatología entre 1982 y 1995. A las mujeres con riesgo de isoinmunización se les aplicó 150 µg de g-globulina anti-D, con fines preventivos. Resultados. En el periodo de estudio ingresaron 4 857 mujeres Rh ne-gativo (4.85 por ciento del total de mujeres), de las cuales 629 (13.0 por ciento), presentaron isoinmunización al RhD. De estas últimas, 542 (86.2 por ciento) ya se encontraron isoinmu-nizadas desde antes de su ingreso al Instituto. En 22 casos (3.5 por ciento), la isoinmunización ocurrió a pesar de que recibie-ron la dosis adecuada de g-globulina anti D. De las 2 605 pacientes (53.6 por ciento) sometidas a prevención, a 2 039 se les aplicó una sola dosis, y a 475, hasta dos dosis. En 22 casos se documentó la falla de la prevención; sin embargo, en cua-tro de ellos, se registraron embarazos múltiples, y los restantes 18 presentaron patología obstétrica asociada. Conclusiones. Mediante este programa de prevención, consistente en administrar 150 µg de g-globulina anti-D por dosis, es posible reducir la iso-inmunización a menos de un caso por cada 1 000 mujeres. Los fracasos en la prevención de la isoinmunización se asociaron a condiciones obstétricas agregadas y al incumplimiento de las guías o lineamientos del programa. El texto completo en inglés de este artículo está disponible en: http://www.insp.mx/salud/index.html

Cambios en los valores plasmáticos de interleucina 1-ß y TNF-alfa al nacimiento / Changes on plasmatic values of interleukine 1ß and NTF-alpha in healthy newborn

Objetivo. Describir los cambios durante el periodo de transición del TNF- a e IL-1b en neonatos sanos, bajo efecto de la vía de nacimiento. Material y métodos. Se seleccionaron aleatoriamente a recién nacidos a término, con bajo riesgo perinatal. Se cuantificaron las concentraciones plasmáticas de TNF- a e IL-1b, al nacimiento, a la hora y 24 horas de vida. Resultados. Se incluyeron 110 neonatos, se obtuvieron concentraciones de TNF- a e UL-1b, cuantificados al nacimiento, a la hora de vida y a las 24 horas de vida; fueron de 7.4, 8.7 y 9.3 pg/mL y de 1.1, 1.3 y 0.7 pg/mL, respectivamente. En el TNF- a, se encontró diferencia significativa al nacimiento (12.9 vs 4.4 pg/mL) y la hora de vida (14.7 vs 5.6 pg/mL), con valores más elevados en los nacidos por vía vaginal (p <0.01). No hubo el mismo efecto para la IL-1b. No hubo influencia estadística en relación con sexo, peso al nacer o edad gestacional. Conclusiones. El TNF- a pero no la IL-1b, presentó cambios estadísticos asociados al parto vaginal.

Cambios en los valores de hemoglobina y la reserva de Fe posterior a la transfusión durante la anemia del prematuro / Hemoglobin and iron stores after blood transfusion in infants with anemia of prematurity

Introducción. Se describen los cambios en los valores de hemoglobina (Hb) y reserva de hierro (Fe) en prematuros evaluados durante la anemia del prematuro, en relación a la transfusión de concentrado eritrocitario.Material y métodos. Se estudiaron neonatos con peso igual o menor de 1 500 g y edad gestacional de 32-34 semanas, divididos en grupo 1 (no transfundidos) y grupo 2 (transfundidos). Se determinaron los niveles de Hb y ferritina sérica (FS) en muestras de sangre venosa obtenidas al nacimiento y a las 11 semanas de edad.Resultados. Se incluyeron a 78 lactantes, 63 casos para el grupo 1 y 15 para el grupo 2. Entre ambos grupos, al nacimiento y 11 semanas, mostraron valores de Hb de 17.1 y 16.5 g/dL y FS de 273 y 212 µg/L. A las 11 semanas de vida, Hb de 11.1 y 11.3 g/dL (P no significativa) y de FS 138 vs 252 µg/L (P = 0.001). No hubo diferencia en los casos con deficiencia o sobrecarga de Fe. Entre las edades de corte la caída de Hb fue similar (6.0 y 5.2 g/dL), que representó 0.5 g/dL por semana y la FS disminuyó 134 µg/L y aumentó 40 µg/L para cada grupo.Conclusiones. Al final de la anemia del prematuro, la transfusión de eritrocitos aumenta la reserva corporal de Fe, aunque sin diferencia significativa en los valores de Hb en estos lactantes.