Albendazole metabolites excretion in human saliva as a biomarker to assess treatment compliance in mass drug administration (MDA) anthelmintic programs.

Soil-transmitted-helminth (STH) infections continue to be a persistent global public health problem. Control strategies for STH have been based on the use of mass drug administration (MDA). Coverage and compliance assessment is critical to understanding the true effectiveness of albendazole (ABZ) in those MDA programs. The aims of this work were to characterize the pattern of albendazole and metabolites excretion in human saliva, and to develop a saliva-based biomarker (HPLC drug/metabolite detection) useful to accurately estimate the coverage/compliance in MDA campaigns. The study subjects were 12 healthy volunteers treated with a single oral dose of ABZ (400 mg). Saliva and blood (dried blood spot, DBS) samples were taken previously and between 2 and 72 h post-treatment. The samples were analyzed by HPLC with UV detection, C18 reversed-phase column. ABZ sulphoxide was the main analyte recovered up to 72 h p.t. in blood and saliva. The concentration profiles measured in the blood (DBS samples) were higher (P < 0.05) than those in saliva, however, this ABZ-metabolite was recovered longer in saliva. The in vivo measurement of drugs/metabolites in saliva samples from ABZ-treated volunteers offers strong scientific evidence to support the use of saliva as a valid biological sample for assessing compliance in MDA programs.

Assessment of Diet-Related Changes on Albendazole Absorption, Systemic Exposure, and Pattern of Urinary Excretion in Treated Human Volunteers.

Soil-transmitted-helminth (STH) infections are a persistent global public health problem. Control strategies for STH have been based on the use of mass drug administration (MDA) mainly targeting preschool- and school-aged-children, although there is increasing interest in expanding treatment to include adults and others through community-wide MDA. Coverage assessment is critical to understanding the real effectiveness of albendazole (ALB) treatment in those MDA programs. The work described here aims to (i) evaluate the effect of type of diet (a heavy or light meal) and fasting before ALB treatment on the systemic disposition of ALB and its metabolites in treated human volunteers and (ii) evaluate the potential feasibility of detecting albendazole metabolites in urine. The data reported here demonstrate that the systemic availability of the active ALB-sulfoxide (ALBSO) metabolite was enhanced more than 2-fold after food ingestion (a heavy or light meal). ALB dissolution improvement related to the ingestion of food may modify the amount of drug/metabolites reaching the parasite, affecting drug efficacy and the overall success of MDA strategies. The measurement in urine samples of the amino-ALB-sulfone (NHALBSO2) derivative and ALBSO for up to 96 h suggests that it may be feasible to develop a noninvasive tool to evaluate compliance/adherence to ALB treatment.

Immunochemical and Physical Quantitation of Grass and Olive Pollen Allergens: Correlation With Asthma Admissions in Cáceres, Spain.

BACKGROUND AND OBJECTIVES: The association between pollen counts and allergen levels in the air is controversial. Objectives: The aims of the study were to quantify total and major allergen levels of Phleum pratense and Olea europaea and to analyze their correlation with grass and olive pollen counts and the number of asthma attacks attended at Complejo Hospitalario Universitario, Cáceres, Spain. MATERIAL AND METHODS: A volumetric air sampler and a Burkard spore trap were used for pollen and aeroallergen collection during April- June 2011. Filters were extracted, and major allergens were quantified using enzyme-linked immunosorbent assay. RESULTS: May was the main grass pollination period, with a maximum peak of 1362 grains/m3 (May 13). The main pollination period for olive was April 30-May 20, with a maximum peak of 851 grains/m3 (May 11). A moderate correlation was observed between asthma exacerbations and grass pollen counts or Phleum total allergen levels; this became stronger when a 3-day offset was introduced. A significant association was observed between asthma exacerbations and total olive allergen or olive pollen grain levels when a 1-day offset was introduced. The maximum correlation (moderate-high) was observed 4 days and 6 days away from the maximum olive pollen peak and the maximum Ole e 1 peak level, respectively. CONCLUSIONS: This study reveals a significant correlation between grass and olive pollination and an increase in the number of visits to the emergency room for asthma attacks. The aerobiological pattern of allergen levels in the air is similar to that of pollen counts during the grass and olive pollination periods.

Upregulation of TET activity with ascorbic acid induces epigenetic modulation of lymphoma cells.

The Ten Eleven Translocation (TET) enzymes have been found to be mutated in both diffuse large B-cell (DLBCL) and peripheral T-cell (PTCL) lymphomas resulting in DNA hypermethylation. Recent studies in embryonal stem cells showed that ascorbic acid (AA) is a cofactor for TET with a binding site at the catalytic domain, and enhances TET activity. We hypothesized that AA could potentially enhance TET activity in lymphoma cells to cause DNA demethylation, reactivate expression of tumor suppressor genes and enhance chemosensitivity. We demonstrate in vitro that AA treatment of DLBCL and PTCL cells using AA concentrations achievable intravenously increased TET activity leading to DNA demethylation. This epigenetic effect is independent of hydrogen peroxide. AA treatment increased the expression of SMAD1, a tumor suppressor gene known to be suppressed by methylation, and increased chemosensitivity of lymphoma cells. Twenty-nine percent (10/34) of unselected lymphoma patients had plasma AA levels that were deficient suggesting an additional clinical mechanism of TET hypofunction. These data indicate that AA has the potential to modify TET function in lymphoma and enhance chemosensitivity. In addition, the AA deficiency seen in some patients may further impair TET function and contribute to resistance. Clinical trials testing intravenous AA with chemotherapy are warranted.

Analysis of the Structure and Function of FOX-4 Cephamycinase.

Class C ß-lactamases poorly hydrolyze cephamycins (e.g., cefoxitin, cefotetan, and moxalactam). In the past 2 decades, a new family of plasmid-based AmpC ß-lactamases conferring resistance to cefoxitin, the FOX family, has grown to include nine unique members descended from the Aeromonas caviae chromosomal AmpC. To understand the basis for the unique cephamycinase activity in the FOX family, we determined the first X-ray crystal structures of FOX-4, apo enzyme and the acyl-enzyme with its namesake compound, cefoxitin, using the Y150F deacylation-deficient variant. Notably, recombinant expression of N-terminally tagged FOX-4 also yielded an inactive adenylylated enzyme form not previously observed in ß-lactamases. The posttranslational modification (PTM), which occurs on the active site Ser64, would not seem to provide a selective advantage, yet might present an opportunity for the design of novel antibacterial drugs. Substantial ligand-induced changes in the enzyme are seen in the acyl-enzyme complex, particularly the R2 loop and helix H10 (P289 to N297), with movement of F293 by 10.3 Å. Taken together, this study provides the first picture of this highly proficient class C cephamycinase, uncovers a novel PTM, and suggests a possible cephamycin resistance mechanism involving repositioning of the substrate due to the presence of S153P, N289P, and N346I substitutions in the ligand binding pocket.

Robustness to mechanical perturbations of center-launching technique for transparent board-to-board and data server interconnects.

Center-launching technique appears as a promising method to allow single-mode propagation in multi-mode fibers, guaranteeing full transparency to the transmitted optical signal also for applications in board-to-board and data server interconnects. In this paper we show that this technique is robust to mechanical perturbations up to about 1 kHz, demonstrating that the vibrations do not affect the transmission performances. Different experimental configurations are tested in order to exclude multimode propagation and to confirm the only fundamental mode propagation. Finally, a theoretical discussion comments the experimental results.

Sandfly saliva of Lutzomyia ovallesi (Diptera: Psychodidae) as a possible marker for the transmission of Leishmania in Venezuela Andes region.

BACKGROUND & OBJECTIVES: The saliva of the Phlebotominae is highly immunogenic to the vertebrate host and is a determining factor in the Leishmania infection. The aim of this work was to study the saliva of Lutzomyia ovallesi as a possible risk marker for the transmission of Leishmania. METHODS: Two populations of L. ovallesi from different geographical areas and subjected to different environmental conditions were compared by geometric morphometry of the wings, by protein profile analysis of salivary glands and by assessing the presence of anti-saliva protein in human sera confronted with laboratory L. ovallesi saliva. RESULTS: The results showed differences in the isometric size and structure of the wings but no allometric effects. Protein profiles of salivary glands of both the L. ovallesi populations studied were found to be similar, based on 11 protein bands with molecular weights ranging from 16 to 99 kDa. Anti-saliva antibodies were present in human sera, but human sera infected and uninfected with leishmaniasis could not be differentiated. INTERPRETATION & CONCLUSION: We conclude that the saliva of laboratory-reared L. ovallesi is representative of that of the wild population. It is suggested to study the presence of anti-saliva antibodies in other species of sandflies and mosquitoes.

Sistema de actividades metodológicas con enfoque de competencia didáctica para profesores de Farmacología I, de la carrera de Medicina: Matanzas / System of methodological activities with an approach of didactical competence for professors of Pharmacology I in the career of Medicine: in Matanzas

Se asumió como enfoque el de las competencias docentes del profesor universitario en el contexto de la universalización. Se utilizaron métodos teóricos, que posibilitaron el estudio en relación con la concepción de la competencia didáctica que se considera básica en los profesores, para sustentar la propuesta diseñada; los métodos empíricos, permitieron recoger información para fundamentar los resultados finales de la investigación. Como procedimientos estadísticos se tuvo en cuenta la estadística descriptiva con frecuencias y porcentajes. El resultado principal de la investigación fue el diseño de un sistema de actividades metodológicas con enfoque de competencia didáctica. Se consideró factible y pertinente la propuesta diseñada, como una de las vías para el desarrollo de competencia didáctica que redunde en el perfeccionamiento del desempeño docente de los profesores universitarios

We assumed as our approach the teaching competences of the university professor in the context of the universalization. We used theoretic methods, allowing the study in relation with the conception of the didactic competence, considered basic in professors, to support de designed proposition. The empirical methods allowed gathering information to sustain the final results of the investigation. As statistical procedure we took into account the descriptive statistics with frequencies and percentages. The main result of the investigation was the design of a system of methodological activities with a didactic competence approach. The designed proposal was considered feasible and pertinent, as a way to develop the didactical competence leading to the improvement of the teaching performance of the university professors

La video clase y la multimedia como medio de enseñanza-aprendizaje de los contenidos morfológicos / The videoconference and multimedia as teaching-learning means for morphologic contents

En la última década, la videoconferencia y la multimedia están siendo percibidas como los medios con más posibilidades didácticas en contextos de enseñanza superior. En sus inicios, la videoconferencia suplió las limitaciones espacio-temporales de la educación a distancia. Progresivamente, este medio fue encontrando un reconocimiento en instituciones convencionales de enseñanza propiciado por el desarrollo de experiencias de calidad basadas en la incorporación de recursos externos al aula. Desde una perspectiva instrumentalista, los medios son soportes materiales de información que deben reflejar la realidad de la forma más perfecta posible. Responden a un modelo estándar de alumno y a una cultura escolar homogénea, y se consideran por sí mismos instrumentos generadores de aprendizajes. Las características del diseño didáctico y computacional de un entorno hipermedia para aprender se intenta conectar a la concepción de promover el aprendizaje utilizando la computadora. El entorno consiste en un conjunto de actividades basadas en andamiajes tendientes a inducir la construcción de significados mediante la activa participación del estudiante.

During the last decade, the videoconference and multimedia are being perceived as the means with more didactic possibilities in the context of the high education. At the beginning, the videoconference helped avoiding limitations of the distance learning. Progressively, this means began to be recognized in conventional teaching institutions because of the development of qualitative experiences based on the usage of out-class resources. From an instrumentalist perspective, the means are material supports of the information that should express the reality in the most possible perfect way. They answer to a standard model of students and to a homogeneous school culture, and are considered, by themselves, as instruments generating learning. The characteristics of a computational and didactic design of a hypermedia setting to learn are intended to be linked to a conception of promoting learning using computers. The setting is a group of activities based on structures tending to induce the construction of meanings through the active participation of the student.

A transition-state analogue reduces protein dynamics in hypoxanthine-guanine phosphoribosyltransferase.

Hypoxanthine-guanine phosphoribosyltransferase (HGPRT) is the key enzyme in purine base salvage in humans and in purine auxotrophs, including Plasmodium falciparum, the leading cause of malaria. Hydrogen/deuterium (H/D) exchange into amide bonds, quantitated by on-line HPLC and mass spectrometry, has been used to compare the dynamic and conformational properties of human HGPRT alone, the HGPRT-GMP-Mg(2+) complex, the HGPRT-IMP-MgPPi <==> HGPRT-Hx-MgPRPP equilibrating mixture, and the transition-state analogue complex HGPRT-ImmGP-MgPPi. The rate and extent of H/D exchange of 26 peptic peptides, spanning 91% of the primary structure, have been monitored. Human HGPRT has 207 amide H/D exchange sites. After 1 h in D2O, HGPRT alone exchanges 160, HGPRT-GMP-Mg(2+) exchanges 154, the equilibrium complex exchanges 139, and the transition-state analogue complex exchanges 126 of these amide protons. H/D exchange rates are correlated with structure for peptides in (1) catalytic site loops, (2) a connected peptide of the subunit interface of the tetramer, and (3) a loop buried in the catalytic site. Structural properties related to H/D exchange are defined from crystallographic studies of the HGPRT-GMP-Mg(2+) and HGPRT-ImmGP-MgPPi complexes. Transition-state analogue binding strengthens the interaction between subunits and tightens the catalytic site loops. The solvent exchange dynamics in specific peptides correlates with hydrogen bond patterns, solvent access, crystallographic B-factors, and ligand exchange rates. Solvent exchange reveals loop dynamics in the free enzyme, Michaelis complexes, and the complex with the bound transition-state analogue. Proton transfer paths, rather than dynamic motion, are required to explain exchange into a buried catalytic site peptide in the complex with the bound transition-state analogue.

Identification by mass spectrometry of a new alpha-tubulin isotype expressed in human breast and lung carcinoma cell lines.

The extensive C-terminal molecular heterogeneity of alpha- and beta-tubulin is a consequence of multiple isotypes, the products of distinct genes, that undergo several posttranslational modifications. These include polyglutamylation and polyglycylation of both subunits, reversible tyrosination and removal of the penultimate glutamate from alpha-tubulin, and phosphorylation of the beta III isotype. A mass spectrometry-based method has been developed for the analysis of the C-terminal diversity of tubulin from human cell lines. Total cell extracts are resolved by SDS--PAGE and transferred to nitrocellulose, and the region of the blot corresponding to tubulin (approximately 50 kDa) was excised and digested with CNBr to release the highly divergent C-terminal tubulin fragments. The masses of the human alpha- and beta-tubulin CNBr-derived C-terminal peptides are all in the 1500--4000 Da mass range and can be analyzed directly by MALDI-TOF mass spectrometry in the negative ion mode without significant interference from other released peptides. In this study, the tubulin isotype diversity in MDA-MB-231, a human breast carcinoma cell line, and A549, a human non-small lung cancer cell line, is reported. The major tubulin isotypes present in both cell lines are k-alpha 1 and beta 1. Importantly, we report a previously unknown alpha isotype present at significant levels in both cell lines. Moreover, the degree of posttranslational modifications to all isotypes was limited. Glu-tubulin, in which the C-terminal tyrosine of alpha-tubulin is removed, was not detected. In contrast to mammalian neuronal tubulin which exhibits extensive polyglutamylation, only low-level monoglutamylation of the k-alpha 1 and beta 1 isotypes was observed in these two human cell lines.

Primary structure of Noetia ponderosa hemoglobins: functional correlates.

Homo- and heterodimeric hemoglobins have been isolated from the red cells of the arcid clam Noetia ponderosa (Np). These hemoglobins bind oxygen cooperatively. An extensively studied dimeric hemoglobin from another arcid clam, Scapaharca inaequivalvis, exhibits a molecular mechanism for cooperative ligand binding that is radically different from tetrameric vertebrate hemoglobins. In this study, the two chains found in both Noetia hemoglobins are sequenced and compared to the hemoglobins of the related clam S. inaequivalvis to determine whether Noetia hemoglobins have the structural basis for the same unusual mechanism for cooperative ligand binding and to inquire about the structural basis of absence of tetramers. Although the Noetia sequences are homologous to the Scapharca sequences, critical differences exist. The lack of tetramerization of Np subunits is most likely related to the absence of critical residues in the A and G helices that stabilize the interdimer contact seen in the Scapharca Hb tetramer. The lower affinity of the homodimer (Np-I), but particularly the heterodimer (Np-II) with respect to the homodimer and heterotetramer of Scapharca, can be due to (i) changes in the proximal heme environment and (ii) changes in the dimer interface. Interactions between Asn 100 and the heme of the other subunit are altered in Np-II due to the substitution of this residue by methionine, possibly causing the reduced O(2) affinity of the heterodimer of Noetia. (iii) Sequence changes in the E and F helices present in Np-I and Np-II could also contribute to the effect through interfacial changes. In particular, the substitution of Val for Thr in position 72 is expected to have a substantial influence on the interface. We conclude that Np dimers have the structural basis for a direct heme-heme interaction mechanism for cooperativity, as in Scapharca, but there are enough sequence changes to suggest that the pathway of interaction might be somewhat different.

Immucillin-H binding to purine nucleoside phosphorylase reduces dynamic solvent exchange.

The rate and extent of hydrogen/deuterium (H/D) exchange into purine nucleoside phosphorylase (PNP) was monitored by electrospray ionization mass spectrometry (ESI-MS) to probe protein conformational and dynamic changes induced by a substrate analogue, products, and a transition state analogue. The genetic deficiency of PNP in humans is associated with severe T-cell immunodeficiency, while B-cell immunity remains functional. Inhibitors of PNP have been proposed for treatment of T-cell leukemia, to suppress the graft-vs.-host response, or to counter type IV autoimmune diseases without destroying humoral immunity. Calf spleen PNP is a homotrimer of polypeptide chains with 284 amino residues, molecular weight 31,541. Immucillin-H inhibits PNP with a Kd of 23 pM when only one of the three catalytic sites is occupied. Deuterium exchange occurs at 167 slow-exchange sites in 2 h when no catalytic site ligands are present. The substrate analogue and product prevented H/D exchange at 10 of the sites. Immucillin-H protected 32 protons from exchange at full saturation. When one of the three subunits of the homotrimer is filled with immucillin-H, and 27 protons are protected from exchange in all three subunits. Deuterium incorporation in peptides from residues 132-152 decreased in all complexes of PNP. The rate and/or extent of deuterium incorporation in peptides from residues 29-49, 50-70, 81-98, and 112-124 decreased only in the complex with the transition state analogue. The peptide-specific H/D exchange demonstrates that (1) the enzyme is most compact in the complex with immucillin-H, and (2) filling a single catalytic site of the trimer reduces H/D exchange in the same peptides in adjacent subunits. The peptides most highly influenced by the inhibitor surround the catalytic site, providing evidence for reduced protein dynamic motion caused by the transition state analogue.

Development of Leishmania (Viannia) braziliensis and Leishmania (Leishmania) amazonensis in the sand fly Lutzomyia migonei (Diptera: Psychodidae).

Development of Leishmania braziliensis (Vianna) and Leishmania amazonensis (Lainson and Shaw) in the sand fly Lutzomyia migonei (França) was compared by studying the parasite microhabitats in the alimentary tract, the sequence of parasite morphological changes leading to the metacyclogenesis process, and the parasite transmission to the vertebrate susceptible host. Although the infections by the 2 Leishmania species were initiated with the same number of amastigotes, Le. amazonensis developed a higher population. Infections with Le. braziliensis were typically peripylarian and those with Le. amazonensis suprapylarian but with an unusual invasion of an organ other than the gut, the Malpighian tubules. The life cycle of the 2 parasites within the sand fly vector included the development of all promastigote forms: procyclics, haptomonads, nectomonads, paramastigotes and infective metacyclics, the last of which are uniquely adapted for transmission to the vertebrate hosts. Appearance of metacyclics coincided with the presence of large number of procyclics and haptomonads, low numbers of nectomonads and the appearance of paramastigotes. In both type of infections, there was a high mortality of the promastigotes inside the bloodmeal during digestion but once infection became established metacyclic forms appeared. Although the numbers of metacyclics that developed in sand flies were low for both parasites they were able to transmit the infection to vertebrates, a key event in the vector competence. We suggest that L. migonei is a true biological host and a possible vector of the 2 Leishmania species, which coexist in extensive geographic areas.

Collagenase degrades collagen in vivo in the ischemic heart.

Previously, we showed that ischemic rat heart contains an activated procollagenase capable of degrading collagen in vitro. We now demonstrate that the collagen resident in such hearts (in vivo) also becomes degraded, producing characteristic fragments implicating the action of an activated collagenase. The evidence is the appearance of amino-terminal dansyl-Ile (+dansyl-Leu) residues in pepsin digests of re-oxygenated rat hearts and immunoblots showing 3/4 length (alphaA) fragments from type I collagen. Also, in ischemic rat myocardium, alphaA(I) and alphaA(III) fragments were detected in pepsin digests. The time periods required for the cleavage and degradation of collagen suggest the participation of a procollagenase that becomes activated. Results demonstrate for the first time that an interstitial collagenase in such hearts initiates in vivo degradation of types I and III collagens.

Human testicular glutathione S-transferases: insights into tissue-specific expression of the diverse subunit classes.

Cytosolic glutathione S-transferase (GST) subunits from human testis were resolved by HPLC and unambiguously identified by combined use of peptide sequence-specific antisera and electrospray ionization mass spectrometry (ESI MS). Allelic variants of hGSTP1, hGSTM1 and hGSTA2 were distinguished on the basis of observed differences in their molecular masses. Relative amounts of the multiple different subunit types in various human tissues were determined from HPLC profiles. From this type of analysis, tissues from hGSTM1 null allele individuals were readily discerned at the protein level; liver was the only tissue in which the hGSTM1 subunit was the major mu-class GST. hGSTM4 and hGSTM5 subunits were found at very low levels in all tissues examined. By far the tissue richest in the unique hGSTM3 subunit was testis, although brain also has significant levels.

Identification of the covalent flavin attachment site in sarcosine oxidase.

Sarcosine oxidase from Corynebacterium sp. P-1 is a heterotetrameric enzyme (alphabetagammadelta) that contains two noncovalently bound coenzymes (FAD, NAD+) and covalently bound FMN [8alpha-(N3-histidyl)FMN] which is attached to the beta subunit. Chlumsky et al. [(1995) J. Biol. Chem. 270, 18252-18259] tentatively identified His175 as the covalent FMN attachment site in the beta subunit, based on an alignment of the sequence of C. sp. P-1 beta subunit with a highly homologous flavin-containing peptide from another corynebacterial sarcosine oxidase (C. sp. U-96). To test this hypothesis, His175 in the C. sp. P-1 beta subunit was mutated to an alanine. Unexpectedly, the mutant enzyme was found to contain 1 mol of covalently bound flavin and to exhibit catalytic activity similar to wild-type enzyme. Covalent flavin-containing peptides were isolated from wild-type and mutant enzymes and analyzed by electrospray mass spectrometry. The mass observed for the mutant peptide (1152.4 Da) matched that predicted for an FMN-containing hexapeptide, corresponding to residues 173-178 (1152.1 Da). In the mutant, this region (HDAVAW) contains a single histidine (His173) which must be the covalent flavin attachment site. The mass observed for the wild-type peptide (1218.6 Da) matched that predicted for an FMN-containing hexapeptide, also corresponding to residues 173-178 in the beta subunit (1218.2 Da). This region in the wild-type enzyme includes two histidine residues (HDHVAW). Attempts to sequence the wild-type or mutant peptides by automated Edman degradation were unsuccessful. Instead, the peptide sequences were investigated by collisional-activated dissociation (CAD) and tandem mass spectrometry. The CAD mass spectral data with the mutant peptide confirmed the sequence deduced based on the mass of the intact peptide. The CAD mass spectral results with the wild-type peptide showed that FMN was covalently attached to the N-terminal histidine in the hexapeptide, which corresponds to His173 in the beta subunit.

Subunit diversity and tissue distribution of human glutathione S-transferases: interpretations based on electrospray ionization-MS and peptide sequence-specific antisera.

Uncertainties about the composition and identities of glutathione S-transferases (GSTs) in human tissue have impeded studies on their biological functions. A rigorous protocol has therefore been developed to characterize the human proteins. Cytosolic GST subunits were resolved by reverse-phase HPLC methods, individual components were assigned to Alpha, Mu and Pi classes on the basis of their immunoreactivities, and peptide-sequence-specific antisera were used to distinguish among five different Mu-class subunits (GSTM1-GSTM5). Each subunit type was characterized and identified unambiguously by electrospray ionization-MS. Acetylation of N-terminal residues in the GSTA1, GSTA2, GSTM3 and GSTM4 subunits were the only natural post-translational modifications detected. The unique structure of GSTM3, with N- and C-terminal peptide extensions predicted from cDNA sequences, was confirmed. Only testis and brain were rich sources of GSTM3 subunits. Subunit profiles were distinct and characteristic of the particular tissue type, and this tissue specificity in GST expression was evident even in organs from different individuals. For instance, livers had relatively simple GST compositions, consisting of a preponderance of Alpha-class subunits and GSTM1 (when present). By contrast, representation of most subunit types was a characteristic feature of testis, which had the highest levels of GSTs. GSTM4 and GSTM5 subunits, here identified for the first time in human tissue extracts, were minor components, with GSTM5 found only in brain, lung and testis. Specimens devoid of GSTM1 subunits, particularly those from null-genotype individuals, were readily discerned at the protein level. Liver was the only rich source of the GSTM1 subunit (although it also constituted a major fraction of adrenal GSTs), and so the functional consequences of the GSTM1 gene deletion are likely to vary in extrahepatic tissues.