Upper respiratory tract viral load quantification in COVID-19 patients at hospital admission and its association with disease severity.

INTRODUCTION: There are aspects of COVID-19 pathogenesis that are still unknown. OBJECTIVE: To determine the relationship between severity, mortality and viral replication in patients with COVID-19. METHODS: Clinical characteristics, severity and mortality of 203 patients hospitalized for COVID-19 were analyzed and correlated with viral load (VL) and threshold cycle (TC) at admission; nasopharyngeal swab was obtained. RESULTS: Mean VLs in surviving patients with mild to moderate, moderate to severe and severe disease were the following: 6.8 x 106, 7.6 x 107 and 1.0 x 109, respectively; and in patients with critical disease who died, VL was 1.70 x 109. TCs were 26.06, 24.07, 22.66 and 21.78 for the same groups. In those who died, a higher mean VL was observed at admission in comparison with those who survived (1.7 x 109 vs. 9.84 x 106; p < 0.001). A significant correlation was observed between VL, severity and death (r = 0.254, p < 0.045 and r = 0.21, p < 0.015). High VL was associated with increased in-hospital mortality in comparison with low VL (OR = 2.926, p < 0.017). CONCLUSION: SARS-CoV-2 VL determined at hospital admission might classify risk simultaneously with other factors described in COVID-19.

INTRODUCCIÓN: Aún se desconocen aspectos de la patogenia de COVID-19. OBJETIVO: Determinar la relación entre gravedad, mortalidad y replicación viral en pacientes con COVID-19. MÉTODOS: Se analizaron características clínicas, gravedad de la enfermedad y mortalidad de 203 pacientes hospitalizados por COVID-19 y se correlacionaron con carga viral (CV) y ciclo umbral (Ct) al ingreso; se tomó hisopado nasofaríngeo. RESULTADOS: Las CV medias en los pacientes sobrevivientes fueron las siguientes ante enfermedad leve a moderada, moderada a grave y grave: 6.8 × 106, 7.6 × 107 y 1.0 × 109; y en los pacientes con enfermedad crítica que fallecieron, la CV fue de 1.70 × 109. Los Ct fueron 26.06, 24.07, 22.66 y 21.78 para esos mismos grupos. En quienes fallecieron se observó mayor CV media al ingreso en comparación con quienes sobrevivieron (1.7 × 109 versus 9.84 × 106), p < 0.001. Se evidenció correlación significativa entre CV, gravedad y muerte (r = 0.254, p < 0.045 y r = 0.21, p < 0.015). La CV alta se asoció a mayor mortalidad intrahospitalaria en comparación con la CV baja (RM = 2.926, p < 0.017). CONCLUSIÓN: La CV de SARS-CoV-2 determinada al ingreso hospitalario podría calificar el riesgo simultáneamente con otros factores descritos en COVID-19.

Activation-dependent mitochondrial translocation of Foxp3 in human hepatocytes.

Foxp3 is considered to be the master regulator for the development and function of regulatory T cells (Treg). Recently Foxp3, has been detected in extra lymphoid tissue, and in hepatocytes and has been associated with hepatocellular carcinoma (HCC), although its role has not been defined. Since it is expected that there is a relationship between protein localization, activity and cellular function, the aim of this study was to explore the subcellular localization of Foxp3 in resting and stimulated human hepatocytes. Foxp3 expression was measured by flow cytometry, subcellular fractioning, and immunofluorescence, and this data was used to track the shuttling of Foxp3 in different subcellular compartments in hepatocytes (HepG2 cell line), stimulated by using the PKC activators (PMA), core and preS1/2 antigen from hepatitis B virus (HBV). Our data shows that besides the nuclear location, mitochondrial translocation was detected after stimulation with PMA and at to a lesser extent, with preS1/2. In addition, Foxp3 is localizes at outer mitochondrial membrane. These results suggest a non-canonical role of Foxp3 in the mitochondrial compartment in human hepatocytes, and opens a new field about their role in liver damages during HBV infection.

Identification of Nef-HIV-1 domains involved in p22-phox interaction and superoxide production.

Nef -HIV-1 has been shown to be involved in NADPH complex interaction and superoxide production. The aim of this work was to study the domains involved in the interaction between Nef and p22-phox. Two approaches were used: 1) in silico modelling, to determine the potential binding motifs and design Nef truncated forms and 2) functional assays. The results showed that GFPVT 68-72, FPDW 121-124 and REVLE 179-183 on Nef are critical for p22-phox (RPQIG 142-146 and PGGP 181-184) docking. However, only the region containing FPDW 121-124 on Nef is able to induce superoxide production. Understanding the molecular mechanisms involved in generating oxidative stress during HIV infection, is critical for therapeutic intervention, in order to minimize viral replication and dissemination.

[Effects of occupational exposure to pesticides on semen quality of workers in an agricultural community of Merida state, Venezuela]. / Efectos de la exposición ocupacional a plaguicidas sobre la calidad del semen en trabajadores de una comunidad agrícola del estado Merida, Venezuela.

Numerous studies report adverse effects of pesticides on male reproductive health. The objectives of this study were to investigate whether there is a relationship between occupational exposure to pesticides and semen quality, and to determine whether chronic exposure to pesticides differentially affects semen quality in men of different ages. A comparative study of 64 farmers and 64 control men was performed. The farmers were interviewed to determine their occupational history and particularly, activities that may involve exposure to pesticides. Semen parameters were evaluated and a comparative analysis of semen variables between exposed and control groups, as well as between age groups: 18-29, 30-37 and 38-60 years was done. Significant alterations of some semen parameters in the exposed group were found, such as: decreases in sperm concentration, slow progressive motility and sperm membrane integrity; at the same time, increases in eosin Y positive and sperm DNA fragmentation index. The results obtained by age groups showed significant differences between exposed and control groups for the parameters of membrane integrity, eosin Y positive and sperm DNA fragmentation index, being the exposed group between 18-29 years that showed the highest altered cases of these parameters. Our results prove that occupational pesticide exposure is associated with alterations in sperm quality, creating a risk to farm workers in their reproductive capacity.

Programmed hepatocytes cell death associated with FLIP downregulation in response to extracellular preS1/2.

Chronic hepatitis B virus (HBV) infection involves liver damage resulting in continuous cell injury and death. During HBV infection, hepatocytes exhibit changes in death receptor expression and in their susceptibility to death. These changes are observed not only in infected cells but also in bystander cells. Because excess viral surface protein (HBsAg) is secreted in large amounts as soluble particles containing preS proteins, the role of soluble preS1/2 in hepatocyte (HepG2) death modulation is an important issue to be explored. An increase of cell death induced by preS1/2 was observed. Also, cell death was associated with the down-regulation of FLIP and activation of caspase 8, caspase 9, and BID. Additionally, hepatocytes exhibited a sensitization to death mediated by the Fas receptor. These results, may contribute to understanding the role of envelope proteins (preS1/2) in the pathogenesis of HBV infection.

A deficient translocation of CD3ζ, ZAP-70 and Grb2 to lipid raft, as a hallmark of defective adaptive immune response during chronic hepatitis B infection.

Hepatitis B is considered to be a worldwide public health problem. An immunosuppressor microenvironment has been proposed to contribute to viral persistence during chronic disease. Understanding the intracellular signaling cascade in T-cells from HBV-infected patients, will contribute to unravel the mechanisms that control the development of immune response during hepatitis B. We analyze lipid rafts formation and early activation signals in chronic HBV infected patients, compared to naturally immune subjects (NIS). Patients show: (1) diminished GM1 clustering, (2) A deficient lipid rafts recruitment of CD3ζ/ZAP-70/Grb2, and (3) these proteins do not merge with GM1 within the lipid rafts. Finally, immunoprecipitation assays proved that ZAP-70 does not associate to CD3ζ. These results show for the first time, defects regarding early key events in T-cell activation, in chronically infected HBV patients, which may contribute not only to understand HBV immune tolerance, but to reveal new potential therapeutic targets to control the infection.

Fas and FasL expression in leukocytes from chronic granulomatous disease patients.

Abstract. Chronic Granulomatous Disease (CGD) is a primary immunodeficiency characterized by defects in superoxide (O2-) production, which result from mutations in one of the four NADPH oxidase components, predisposing to bacterial and fungal infections. Besides the O2-defect, it has been described that neutrophils from CGD patients are resistant to cell death, a phenomenon that has been connected to chronic inflammation and predisposition to autoimmune diseases. A diminished expression of Fas and its counterpart FasL, molecules known to play a major role in cell death, has been described in lymphocytes depleted of O2-reactive oxygen species (ROS), suggesting an involvement of ROS in Fas/FasL expression. In this work, Fas and FasL expressions were analyzed in T cells and neutrophils from two CGD families, previously known to harbor two different molecular defects: absence of either p47-phox or p67-phox. We found that T lymphocytes from CGD patients express low levels of Fas and FasL, while a diminished FasL expression was observed on neutrophils from a CGD A470 patient. These defects may contribute to understand altered cell death in CGD patients.

[Analysis of p22-phox and p47-phox subcellular localization and distribution in neutrophils from human immunodeficiency virus (HIV) infected patients]. / Localización y distribución celular de p22-phox y p47-phox en neutrófilos humanos de pacientes infectados con VIH.

INTRODUCTION: During human immunodeficiency virus (HIV) infection a dysfunction of polymorphonuclear (PMN) cells has been described including a progressively altered superoxide production as disease progression. The NADPH oxidase has been described as a major source of superoxide. The neutrophil NADPH oxidase comprises a plasma membrane-bound cytochrome b558 (which is a heterodimer of one p22-phox and one gp91-phox subunit) and cytosolic subunits, namely p47-phox, p67-phox and p40-phox. During neutrophil activation in response to various agonists, the cytosolic subunits translocate to and associate with the cytochrome b558, a process that results in oxidase activation. Therefore, an altered superoxide production could be a consequence of abnormal distribution or translocation of NADPH oxidase components in response to HIV infection. MATERIAL AND METHODS: We used several strategies including: confocal microscopy, subcellular fractionation and sucrose gradients, to analyze the cellular distribution of two of the NADPH oxidase components (p22-phox and p47-phox). RESULTS: We observed that in resting cells, a substantial proportion of p22-phox from HIV positive patients is distributed in regions close to the cytoplasmic membrane, sediment in high density sucrose fractions and is located in the cytoplasmic insoluble fraction. Additionally, a diffuse cytosolic distribution of p47-phox was observed in neutrophils from HIV infected patients. The results demonstrate an inappropriate cell distribution of NADPH-complex in PMN from HIV positive patients.

HIV-1 Nef associates with p22-phox, a component of the NADPH oxidase protein complex.

Altered neutrophil function may contribute to the development of AIDS during the course of HIV infection. It has been described that Nef, a regulatory protein from HIV, can modulate superoxide production in other cells, therefore altered superoxide production in neutrophils from HIV infected patients, could be secondary to a direct effect of Nef on components of the NADPH oxidase complex. In this work, we describe that Nef, was capable of increasing superoxide production in human neutrophils. Furthermore, a specific association between Nef and p22-phox, a membrane component of the NADPH oxidase complex, was found. We propose that this association may reflect a capability of Nef to modulate by direct association, the enzymatic complex responsible for one of the most efficient innate defense mechanisms in phagocytes, contributing to the pathogenesis of the disease.

[Immune tolerance at 50 years from the Nobel Prize in Medicine and Physiology: a perspective as a mechanism of immune response against pathogens]. / La tolerancia inmunológica a 50 años del Premio Nobel en Medicina y Fisiología: una perspectiva como mecanismo de respuesta inmune frente a patógenos.

The immune system has the capability of protection against infectious disease which is accomplished by an enormous repertoire of receptors specifically reactive to foreign antigens, but it is tolerant to self-antigens, establishing biological identity. The ability to discriminate between self and non-self is a central property of the immune system, by using complex network of cellular and molecular mechanisms in order to prevent autoimmunity; this function is called immune tolerance. Thus, the Interaction between immune system and antigens is required for the generation of tolerance and it is critical in different physiological and pathological conditions in order to limit the damage to self tissues. Since Medawar description, who showed that the tolerance is an acquired property playing a central role in the homeostasis, several mechanism has been proposed to explain it. It is accepted today that an important group of antigen specific cells called regulatory T cell, both natural and induced, are critical as a unifying mechanism to maintain self-tolerance. These suppressor lymphocyte subpopulations had shown to play an important role not only by controlling autoimmune disease but also in the pathogenesis of many chronic infectious diseases, either manipulated by the microorganism to escape from the immune system, or induced by the host to reduce inflammatory damage. This review has the intention of updating about modern concepts in immune tolerance mechanisms, with special emphasis played by Treg cell in the tolerance which is unquestionable induced during the course of chronic infections diseases.

[Evaluation of the quality of the human spermatozoon: comparison between spermatic DNA integrity and semen variables]. / Evaluación de la calidad del espermatozoide humano: comparación entre la integridad del ADN espermático y variables del semen.

Semen analysis does not have an absolute predictive value on fertility, however it is a reflection of male fertility potential, which is related to its spermatozoa quality and other semen variables. Great variability in human semen parameters has been demonstrated within a single individual, an observation that could explain why a male with low semen quality can successfully fertilize an egg. Although conventional semen analysis, such as sperm concentration, motility and morphology, provide important information about the clinical status of male fertility, new procedures to predict the sperm functional capability have been developed in the last decade, such as analysis of nuclear DNA integrity, which have improved considerably the clinical diagnosis of male infertility, and increased the knowledge about spermatozoa function. DNA fragmentation consist in interruptions, both in single and double DNA strains, that frequently occur in sperm samples from infertile patients. We have conducted a clinical study in semen samples from patients who have attended the Andrology laboratory of the University of Los Andes, between March 2007 and March 2009. The aim of this study was to compare sperm DNA integrity, analyzed by flow cytometry, with traditional semen parameters. Our results show remarkable correlations between conventional human semen variables and sperm chromatin integrity, contributing to asses an integral evaluation of sperm quality allowing the analysis of its fertilizing potential in clinical studies.

Epidemiological and clinical characteristics of patients with malaria admitted to a hospital in Mérida, Venezuela.

BACKGROUND: Malaria is expanding rapidly across Venezuela, spreading outwards from traditional high transmission regions in the southeast of the country, but the lack of official data make it impossible to understand the reasons for this expansion and to estimate its real magnitude. This study aims to evaluate the epidemiological characteristics driving the re-emergence of malaria in Mérida, a state in the west of Venezuela, where no cases have been reported since 2003, and also to study the clinical presentation of the disease in patients presenting with malaria. METHODS: Thirty-three patients who presented with anemia and fever and with a microscopic diagnosis of malaria were examined and interviewed. Data were collected in standardized forms and analyzed. One-way analysis of variance was used to study differences among patients infected with different parasites. RESULTS: Twenty-two patients were from the Zulia state and eleven were from the Mérida state, mainly from the lowlands south of Lake Maracaibo. Six of these patients traveled to the Bolívar state between 2017 and 2019. Thirteen patients presented with the WHO criteria for severe malaria.Conclusions:Domestic migration to the southeast of Venezuela may have played an important role in the expansion of malaria in previously existing endemic areas of transmission and also in the increase in the number of cases of severe malaria.

Altered T cell costimulation during chronic hepatitis B infection.

T-cell response to hepatitis B virus (HBV) is vigorous, polyclonal and multi-specific in patients with acute hepatitis who ultimately clear the virus, whereas it is narrow and inefficient in patients with chronic disease, where inappropriate early activation events could account for viral persistence. We investigated the induction of activation receptors and cytokine production in response to HBcAg and crosslinking of CD28 molecules, in CD4+ cells from a group of chronically infected patients (CIP) and naturally immune subjects (NIS). We demonstrated that CD4+ cells from CIP did not increase levels of CD40L and CD69 following stimulation with HBcAg alone or associated to CD28 crosslinking, in contrast to subjects that resolved the infection (p<0.01). Furthermore, CD4+ cells from CIP produced elevated levels of IL-10 in response to HBcAg. These results suggest that a predominant inhibitory environment may be responsible for altered T cell costimulation, representing a pathogenic mechanism for viral persistence.

[Impaired activation and costimulation of T CD4+ lymphocytes during chronic hepatitis C infection]. / Déficit en señales de activación y coestimulación en linfocitos T CD4+ de pacientes infectados con el virus de la hepatitis C.

Hepatitis C chronic infection occurs in 80% of the cases and eventually leads to cirrhosis and hepatocellular carcinoma. A deficient adaptive immune response has been described during chronic infection which contributes to viral persistence. This altered T cell response could be associated to deficient costimulation signals during priming of T cells. We have conducted an in vitro study to explore the activation phenomenon of CD4+ T cells focusing on costimulation via the CD28 receptor, associated to stimulation with purified Hepatitis C (HCV) core antigen. Our study involved the induction of CD69, CD25 and CD40L activation receptors, along with detection of intracellular cytokines such as IFN-gamma, TGF-beta and IL-10. Analysis was performed in chronically HCV infected patients, intrafamilial members of HCV-infected patients and healthy individuals. HCV core antigen induced CD40L expression in CD4+ cells from intrafamilial members, in contrast to chronically infected patients and control individuals. Association of CD28 crosslinking increased CD69 and IFN-gamma expression in chronically infected patients, suggesting a detriment in this signaling pathway. Additionally, an increased TGF-beta expression was observed in CD4+ cells from HCV-infected patients, which was corrected by addition of CD28 crosslinking. Our results may contribute to understand the underlying mechanism of T cell tolerance against HCV during chronic infection, and to provide new targets for the designing of therapeutic strategies to control the infection and to offer protective immunity against the virus.

CD40/CD40L expression in leukocytes from chronic granulomatous disease patients.

Chronic granulomatous disease (CGD) is an inherited disorder caused by defects in the NADPH oxidase complex, which generates superoxide, the precursor of hydrogen peroxide (H(2)O(2)) and other reactive oxygen derivatives with microbicidal activity. Because CGD patients are at risk of chronic inflammatory manifestations, including inflammatory bowel disease and autoimmune diseases, and it is not clear whether these pathologies are exclusively secondary to altered superoxide production, or whether distinct immunologic defects are involved, we explored cell proliferation, lymphocyte cell counts, immunoglobulin levels, presence of autoimmune antibodies and expression of costimulatory molecules in leukocytes from CGD patients. We found that CGD patients have a diminished phytohemagglutinin-induced proliferation of blood mononuclear cells. Following stimulation with PMA plus ionomycin, a reduced percentage of CD40L expression in T lymphocytes and a diminished expression of CD40 molecules in neutrophils were observed on leukocytes from these patients. Our results suggest an altered interplay between elements of innate and adaptive immunity in CGD patients, which may be reflected in an increased susceptibility to opportunistic infections.

[Oral mucosa analog allografts in non-consanguineous rats].

INTRODUCTION: Although there are therapeutic options for the treatment of oral mucosa defects, the need for functional, anatomical and aesthetically similar substitutes persists, as well as for solutions to reduce autologous grafts morbidity. OBJECTIVE: To determine clinical and histological compatibility of equivalent oral mucosa allografts generated through tissue engineering in non-consanguineous rats. MATERIALS AND METHODS: We used a sample of oral mucosa from Sprague Dawley rats to obtain a fibroblast culture and a keratinocytes and fibroblasts co-culture. In both cases, we used a commercial collagen membrane as "scaffold". After ten weeks of culture, we grafted the resulting membranes into four Wistar rats. The first phase of the study was the development of the oral mucosa equivalents generated by tissue engineering. Then, we implanted them in immunocompetent Wistar rats, and finallywe evaluated the clinical and histological features of the allografts. RESULTS: In vivo evaluation of mucosal substitutes showed a correct integration of artificial oral mucosa in immunocompetent hosts, with an increase in periodontal biotype and the creation of a zone with increased keratinization. Histologically, the tissue was similar to the control oral mucosa sample with no inflammatory reaction nor clinical or histological rejection signs. CONCLUSION: The equivalent oral mucosa allografts generated by tissue engineering showed clinical and histological compatibility.

Cuantificación de la carga viral en el tracto respiratorio superior en pacientes con COVID-19 al ingreso hospitalario y su asociación con la gravedad de la enfermedad / Upper respiratory tract viral load quantification in COVID-19 patients at hospital admission and its association with disease severity

Resumen Introducción: Aún se desconocen aspectos de la patogenia de COVID-19. Objetivo: Determinar la relación entre gravedad, mortalidad y replicación viral en pacientes con COVID-19. Métodos: Se analizaron características clínicas, gravedad de la enfermedad y mortalidad de 203 pacientes hospitalizados por COVID-19 y se correlacionaron con carga viral (CV) y ciclo umbral (Ct) al ingreso; se tomó hisopado nasofaríngeo. Resultados: Las CV medias en los pacientes sobrevivientes fueron las siguientes ante enfermedad leve a moderada, moderada a grave y grave: 6.8 × 106, 7.6 × 107 y 1.0 × 109; y en los pacientes con enfermedad crítica que fallecieron, la CV fue de 1.70 × 109. Los Ct fueron 26.06, 24.07, 22.66 y 21.78 para esos mismos grupos. En quienes fallecieron se observó mayor CV media al ingreso en comparación con quienes sobrevivieron (1.7 × 109 versus 9.84 × 106), p < 0.001. Se evidenció correlación significativa entre CV, gravedad y muerte (r = 0.254, p < 0.045 y r = 0.21, p < 0.015). La CV alta se asoció a mayor mortalidad intrahospitalaria en comparación con la CV baja (RM = 2.926, p < 0.017). Conclusión: La CV de SARS-CoV-2 determinada al ingreso hospitalario podría calificar el riesgo simultáneamente con otros factores descritos en COVID-19.

Abstract Introduction: There are aspects of COVID-19 pathogenesis that are still unknown. Objective: To determine the relationship between severity, mortality and viral replication in patients with COVID-19. Methods: Clinical characteristics, severity and mortality of 203 patients hospitalized for COVID-19 were analyzed and correlated with viral load (VL) and threshold cycle (TC) at admission; nasopharyngeal swab was obtained. Results: Mean VLs in surviving patients with mild to moderate, moderate to severe and severe disease were the following: 6.8 × 106, 7.6 × 107 and 1.0 × 109, respectively; and in patients with critical disease who died, VL was 1.70 × 109. TCs were 26.06, 24.07, 22.66 and 21.78 for the same groups. In those who died, a higher mean VL was observed at admission in comparison with those who survived (1.7 × 109 vs 9.84 × 106; p < 0.001). A significant correlation was observed between VL, severity and death (r = 0.254, p < 0.045 and r = 0.21, p < 0.015). High VL was associated with increased in-hospital mortality in comparison with low VL (OR = 2.926, p < 0.017). Conclusion: SARS-CoV-2 VL determined at hospital admission might classify risk simultaneously with other factors described in COVID-19.

[Pattern of T cell activation in absence of protective immunity against hepatitis B virus. Review]. / Patrón de activación de linfocitos T en ausencia de respuesta protectora contra el virus dela hepatitis B. Revisión.

Hepatitis B is an important cause of morbidity and mortality around the world. One-third of the world's population has been estimated to be infected with hepatitis B virus (HBV). A significant amount of evidence suggests that both humoral and cellular immune responses are important to eliminate the virus and that, cellular immunity is involved in the pathogenesis of the disease. Vaccination with HBsAg is considered as the main strategy for effective control of the infection and viral transmission. However, approximately 5-10% of immunized individuals fail to elicit detectable specific antibodies and remain at risk for hepatitis B infection. In this work we have reviewed the current status in the pathogenesis of the disease and the mechanisms described to explain nonresponsiveness to the vaccine as well. Since nonresponders to the vaccine are at risk for the infection, a common mechanism to explain the absence or inappropriate immune response to virus components is proposed. Within the suggested model an impaired activation of T lymphocytes against viral antigens, both in nonresponders to vaccination and chronically infected patients, is described. These observations could be consistent with potential differences in the MHC/Ag presentation; therefore contributing to our understanding of the altered T helper response as an underlying mechanism for the lack of protective immunity against VHB.

High prevalence of secondary resistance mutations in Venezuelan HIV-1 isolates.

The genetic variability was studied in HIV-1 from Venezuelan patients with and without treatment, in order to evaluate the presence of polymorphisms and drug resistance mutations. Proviral DNA from peripheral blood mononuclear cells or viral RNA from plasma was extracted from the blood of 30 patients. Two regions from the polymerase gene, protease (Pr) and reverse transcriptase (RT) and one genomic fragment from the envelope (Env) gene were amplified and sequenced. All HIV-1 samples analyzed were classified as subtype B, without evidence of recombination. Although no primary protease mutations were detected, a high frequency of secondary mutations (86%, 19/22), associated to restoration of viral replicative fitness, was observed in strains circulating both in treated and non-treated patients. Resistance mutations to nucleoside RT inhibitors (NRTI) and non-nucleoside RT inhibitors (NNRTI) were detected in 35% (6/17) and 12% (2/17) of the viruses circulating in treated patients, respectively. Resistance mutations were also present in the virus infecting one antiretroviral naive individual (7.7%), suggesting that local screening for resistant mutation in naive patient might be important to minimize therapy failure. Future studies are warranted to assess the role of secondary mutation in the success of viral infection.

[Cytotoxic effect of palladium (II) inclusion compounds in beta-cyclodextrin].

INTRODUCTION: Thiosemicarbazones and palladium (II) complexes have antineoplastic activities with mild side effects, for which they are considered new alternative antineoplastic drugs. However, the IC50 ranges of these complexes vary due to differences in their structure and solubility and their sensitivities for various cellular targets. Beta-cyclodextrin is an additive used to improve the solubility and stability of various drugs for therapeutic use, but the combination of beta-cyclodextrin with palladium (II) complexes and thiosemicarbazones has not been tested yet. OBJECTIVE: To study the cytotoxic effect of palladium (II) inclusion complexes in beta-cyclodextrin. MATERIALS AND METHODS: We tested the cytotoxic activity of palladium complexes combined with beta-cyclodextrin in the breast cancer cell line MCF-7 using a sulforhodamine B assay. RESULTS: We tested the antiproliferative activity of palladium (II) complexes with and without the ligands MePhPzTSC and Ph2PzTSC and with and without beta-cyclodextrin in MCF-7 cells and compared them to that of cisplatin. All combinations showed antiproliferative activity; however, the activity was greater for the combinations that included beta-cyclodextrin: ([Pd (MePhPzTSC) 2] • ß-CD and [Pd (Ph2PzTSC) 2] • ß-CD), at concentrations of 0.14 and 0.49 µM, respectively. The IC50 for this complex was 5-fold lower than that of the ligand-free combinations (1.4 and 2.9 µM, respectively). The IC50 for free palladium (II) complex was 0.571.24 µM and that for cisplatin was 6.87 µM. CONCLUSIONS: Beta-cyclodextrin significantly enhanced the cytotoxic activities of palladium (II) complexes and thiosemicarbazones probably by improving their solubility and bioavailability. The addition of beta-cyclodextrin is a possible strategy for designing new anticancer drugs.