TLR Agonists Modify NK Cell Activation and Increase Its Cytotoxicity in Acute Lymphoblastic Leukemia.

Natural killer (NK) cells play a crucial role in innate immunity, particularly in combating infections and tumors. However, in hematological cancers, NK cells often exhibit impaired functions. Therefore, it is very important to activate its endosomal Toll-like receptors (TLRs) as a potential strategy to restore its antitumor activity. We stimulated NK cells from the peripheral blood mononuclear cells from children with acute lymphoblastic leukemia and NK cells isolated, and the NK cells were stimulated with specific TLR ligands (Poly I:C, Imiquimod, R848, and ODN2006) and we evaluated changes in IFN-γ, CD107a, NKG2D, NKp44 expression, Granzyme B secretion, cytokine/chemokine release, and cytotoxic activity. Results revealed that Poly I:C and Imiquimod enhanced the activation of both immunoregulatory and cytotoxic NK cells, increasing IFN-γ, CD107a, NKG2D, and NKp44 expression. R848 activated immunoregulatory NK cells, while ODN2006 boosted CD107a, NKp44, NKG2D, and IFN-γ secretion in cytotoxic NK cells. R848 also increased the secretion of seven cytokines/chemokines. Importantly, R848 and ODN 2006 significantly improved cytotoxicity against leukemic cells. Overall, TLR stimulation enhances NK cell activation, suggesting TLR8 (R848) and TLR9 (ODN 2006) ligands as promising candidates for antitumor immunotherapy.

[Euthanasia laws in Spain and in the world: Medical aspects]. / Leyes de eutanasia en España y en el mundo: aspectos médicos.

OBJECTIVE: To compare the Spanish law on euthanasia and assisted suicide with those that exist in other countries. DESIGN: Systematic review of the bibliography. DATA SOURCES: Medline/PubMed, EMBASE and the Cochrane Library were searched for studies that contained in their title or abstract the descriptors «euthanasia¼ or «assisted suicide¼ and also «legislation¼ or «law¼, between 2002 and the end of 2020. STUDY SELECTION: The search found 1647 studies and after screening 663 were assessed, of which 30 were included in the review. Studies that only contained opinions or did not provide data on euthanasia/assisted suicide in the countries that have them regulated were rejected. DATA EXTRACTION: We registered the criteria that regulate the acceptance or rejection of a request for euthanasia or assisted suicide in Spain and in the other countries where they are decriminalized. RESULTS: The euthanasia regulations in the world can be grouped into three groups: laws that allow euthanasia and assisted suicide (Netherlands, Belgium, some states of Australia, New Zealand, Spain), those in which the law only allows assisted suicide (USA) and those in which only assisted suicide is admitted and based on court decisions, without specific legislation (Switzerland, Germany). CONCLUSIONS: Although there are differences, the laws that the Spanish euthanasia law most closely resembles are those of the Netherlands and Belgium, so it is foreseeable that the casuistry of euthanasia and its figures in Spain will resemble that of those countries in the future.

High expression of Toll-like receptor 7 is a survival factor in pediatric medulloblastoma.

PURPOSE: Medulloblastoma is an embryonal brain tumor that predominantly occurs in childhood with a wide histological and molecular variability. Our aim was to investigate the expression of Toll-like receptors (TLRs), their association with the infiltration of immune cells and with the histological subgroups, and, also, with the overall survival of patients. METHODS: Fifty-six paraffin-preserved biopsies from children with medulloblastoma of the classic, desmoplastic, and anaplastic subtypes were included. Microarrays of tissues were performed, and the infiltration of T and NK cells was quantified, as well as the expression of TLR7, TLR8, and TLR9. For all statistical analyses, significance was p < 0.05. RESULTS: CD4 + and CD8 + T lymphocytes and NK cells were found infiltrating the tumor. The infiltration of NK and CD4 + cells was greater in the classic and desmoplastic subtypes than in anaplastic. We found an important expression of TLRs in all medulloblastomas, but TLR7 and TLR8 were considerably higher in classic and desmoplastic subtypes than in anaplastic. Importantly, we observed that TLR7 was a prognostic factor for survival. CONCLUSIONS: Medulloblastomas present cellular infiltration and a differential expression of TLRs depending on the histological subtype. TLR7 is a prognostic factor of survival that is dependent on treatment and age.

Granuloma Formation in a Cyba-Deficient Model of Chronic Granulomatous Disease Is Associated with Myeloid Hyperplasia and the Exhaustion of B-Cell Lineage.

Haematopoiesis is a paradigm of cell differentiation because of the wide variety and overwhelming number of mature blood cells produced daily. Under stress conditions, the organism must adapt to a boosted demand for blood cells. Chronic granulomatous disease (CGD) is a genetic disease caused by inactivating mutations that affect the phagocyte oxidase. Besides a defective innate immune system, CGD patients suffer from recurrent hyper-inflammation episodes, circumstances upon which they must face emergency haematopoiesis. The targeting of Cybb and Ncf1 genes have produced CGD animal models that are a useful surrogate when studying the pathophysiology and treatment of this disease. Here, we show that Cyba-/- mice spontaneously develop granuloma and, therefore, constitute a CGD animal model to complement the existing Cybb-/- and Ncf1-/- models. More importantly, we have analysed haematopoiesis in granuloma-bearing Cyba-/- mice. These animals showed a significant loss of weight, developed remarkable splenomegaly, bone marrow myeloid hyperplasia, and signs of anaemia. Haematological analyses showed a sharped decrease of B-cells and a striking development of myeloid cells in all compartments. Collectively, our results show that granuloma inflammatory lesions dramatically change haematopoiesis homeostasis. Consequently, we suggest that besides their defective innate immunity, the alteration of haematopoiesis homeostasis upon granuloma may contribute to the dismal outcome of CGD.

Polymorphisms in IL-10 and TGF-ß gene promoter are associated with lower risk to gastric cancer in a Mexican population.

BACKGROUND: Helicobacter pylori infection is recognized as the main risk factor for gastric cancer (GC), the fifth most common neoplasia worldwide. H. pylori interacts with the immune system, disrupting the cytokine network and inducing chronic inflammation. This work aimed to evaluate the association between single nucleotide polymorphisms (SNPs) in selected cytokine gene promoters and GC. METHODS: The study included 359 subjects, 125 GC patients, 109 intestinal metaplasia (IM) patients and 125 asymptomatic controls. DNA was extracted from white blood cells and nine SNPs in cytokine gene promoters were genotyped using predesigned 5'-endonulease assays. The association of the SNPs with IM and GC was evaluated using multinomial regression models. RESULTS: Both genotypes, TC (OR = 0.51, 95% CI = 0.27-0.98) and TT (OR = 0.42, 95% CI = 0.20-0.91) in the locus - 509 of the TGF-ß promoter were significantly associated with GC. The TT genotype in the locus - 819 of the IL-10 promoter was also significantly associated with GC (OR = 0.37, 95% CI = 0.17-0.81). No significant association was found with SNPs IL-4 -590 T/C (rs1800629), IL-6 -573G/C (rs1800796), IL-10 -592C/A (rs1800872), IL-10 -1082A/G (rs1800896), and, IFN-γ -1615C/T (rs2069705). CONCLUSIONS: SNPs in TGFß (- 509 C/T, rs1800469) and IL-10 (- 819 C/T, rs1800871) promoters were associated with a lower risk for GC in a Mexican population.

Primary Care Physicians Can Comprehensively Manage Patients with Sleep Apnea. A Noninferiority Randomized Controlled Trial.

Rationale: General practitioners play a passive role in obstructive sleep apnea (OSA) management. Simplification of the diagnosis and use of a semiautomatic algorithm for treatment can facilitate the integration of general practitioners, which has cost advantages.Objectives: To determine differences in effectiveness between primary health care area (PHA) and in-laboratory specialized management protocols during 6 months of follow-up.Methods: A multicenter, noninferiority, randomized, controlled trial with two open parallel arms and a cost-effectiveness analysis was performed in six tertiary hospitals in Spain. Sequentially screened patients with an intermediate to high OSA probability were randomized to PHA or in-laboratory management. The PHA arm involved a portable monitor with automatic scoring and semiautomatic therapeutic decision-making. The in-laboratory arm included polysomnography and specialized therapeutic decision-making. Patients in both arms received continuous positive airway pressure treatment or sleep hygiene and dietary treatment alone. The primary outcome measure was the Epworth Sleepiness Scale. Secondary outcomes were health-related quality of life, blood pressure, incidence of cardiovascular events, hospital resource utilization, continuous positive airway pressure adherence, and within-trial costs.Measurements and Main Results: In total, 307 patients were randomized and 303 were included in the intention-to-treat analysis. Based on the Epworth Sleepiness Scale, the PHA protocol was noninferior to the in-laboratory protocol. Secondary outcome variables were similar between the protocols. The cost-effectiveness relationship favored the PHA arm, with a cost difference of €537.8 per patient.Conclusions: PHA management may be an alternative to in-laboratory management for patients with an intermediate to high OSA probability. Given the clear economic advantage of outpatient management, this finding could change established clinical practice.Clinical trial registered with www.clinicaltrials.gov (NCT02141165).

Nested Polymerase Chain Reaction and Cutaneous Tuberculosis.

The role of Mycobacterium tuberculosis in the etiology and pathogenesis of cutaneous tuberculosis is controversial because of the difficulties associated with demonstrating the presence of these mycobacteria in tuberculid cutaneous lesions by routinely available microbiological and histological techniques. In this study, we aimed to demonstrate the presence of M. tuberculosis in cutaneous tuberculosis. Multiple polymerase chain reaction (PCR) followed by nested PCR was used to amplify genomic fragments from 3 different mycobacteria species. DNA was isolated from 30 paraffin-embedded skin biopsies. Samples were selected randomly from patients with a clinical and histopathological diagnosis of the most frequent groups of cutaneous tuberculosis in Mexico as follows: 5 cases of scrofuloderma tuberculosis; 2 cases of lupus vulgaris tuberculosis; and 5 cases of tuberculosis verrucosa cutis. The other cases denominated tuberculids in some countries such as Mexico and included the following: 7 cases of rosacea-like tuberculosis; one case of papulonecrotic tuberculosis; and 10 cases of erythema induratum of Bazin. Four normal skin biopsies were included as controls. M. tuberculosis DNA was amplified successfully by nested PCR in 80% of the samples (24 of the 30 samples) assayed. Mycobacterial DNA was not detected in the normal skin biopsies used as controls. Detection of M. tuberculosis DNA in 80% of cutaneous tuberculosis analyzed implicates this mycobacterium in the pathogenesis of multiple clinical forms of cutaneous tuberculosis.

Suicide in primary headaches in 48 countries: A physician-survey based study.

Aim To investigate the relationship between primary headache types and accomplished or attempted suicide in countries from all world regions. Methods Data were obtained using a questionnaire about suicide due to headache in a face-to-face interview with 203 physicians with expertise in headaches. They came from 48 countries, and from all continents. Results Primary headaches cause one suicide per 1,000,000 population each year (1% of the suicide rate due to all causes). Cluster headache and migraines account for 70-80% of them. Suicide attempts are 10 times more frequent than accomplished suicides. Cluster headache poses more risk than migraine. This risk is not often acknowledged, and is increased if there is previous psychiatric history. More than half of the physicians interviewed think it could be reduced with a more aggressive treatment of headaches. Conclusions Cluster headache and migraine are not always benign, and are the cause of the majority of suicides due to headache.

Giant Prolactinoma in Men: Clinical Features and Therapeutic Outcomes.

The aim of the study was to evaluate the clinical features and long-term therapeutic outcome of giant prolactinoma (gPRLoma) in men and to compare them with those of a group of male patients with non-gPRL macroprolactinomas (non-gPRLomas). A retrospective and multicenter study of gPRLomas in men diagnosed in a 20-year period was performed. Clinical data and treatment outcome were registered. The diagnosis of gPRLoma was established when the maximal tumor diameter was ≥40 mm or the tumor had ≥20 mm of suprasellar extension associated to hyperprolactinemia (PRL>1000 ng/ml). Non-gPRLoma was considered when tumor diameter was ≥ 10 mm and<40 mm associated to hyperprolactinemia (PRL≥200 ng/ml). Twenty-three patients with gPRLoma (age 38.3±13.5 years) followed for at least 3 months (follow-up 87.1±60.5 months, range 3-211 months) were evaluated. A group of 42 patients with non-gPRLoma (age 42±16.6 years; NS; follow-up 89±65.9 months, range 3-222 months; NS) served as a control group. More than half (56.5%) of the gPRLoma patients were younger than 40 years at diagnosis. Visual disturbances were significantly more common in gPRLoma than in non-gPRLoma patients (65.2 vs. 25.6%; p=0.004). Prevalence of hypopituitarism was similar in both groups of patients (73.9% vs. 80.9%; gPRLoma vs non-gPRLoma; NS). Serum PRL concentrations were significantly higher in gPRLoma than in non-gPRLoma patients [median (IR), 3978 ng/ml (1179-9012) vs. 907 ng/ml (428-3119); p<0.001]. Maximum tumor diameter in gPRLomas was 4.8±0.8 cm and 2.4±0.7 cm in non-gPRLoma (p<0.001). All patients were treated with dopamine agonists (DA). Twelve (52.2%) gPRLoma patients and 32 (73.8%) non-gPRLoma patients were treated with DA as monotherapy (p=0.045). Surgery was used in 12 (52.2%) gPRLoma patients and in 12 (28.6%) non-gPRLoma patients (p=0.054). Lastly, radiotherapy was used in 5 (21.7%) gPRLoma patients and in 6 (14.2%) non-gPRLoma patients (NS). At last visit, PRL was similar in both groups of patients [16 ng/ml (4-30) vs. 11 ng/ml (4-25); gPRLomas vs. non-gPRLomas; ns] and tumor size decreased significantly (p<0.001) in both groups of patients. Clinical cure (maintained normoprolactinemia without therapy for>1 year and no radiological evidence of pituitary tumor) was achieved in 2 (8.7%) gPRLoma patients and in 2 (4.8%) non-gPRLoma patients (NS). gPRLomas in men are usually diagnosed at a mean age of 40 years, an age similar to that of non-gPRLomas. The only clinical difference with non-gPRLomas is their greater prevalence of visual disturbances. The therapeutic approaches and tumor outcomes were similar to those obtained in patients with non-gPRLomas. Complete cure in gPRLoma is rare, but similar to that achieved in non-gPRLomas, reached in less than 10% of patients.

Conventional Polysomnography Is Not Necessary for the Management of Most Patients with Suspected Obstructive Sleep Apnea. Noninferiority, Randomized Controlled Trial.

RATIONALE: Home respiratory polygraphy may be a simpler alternative to in-laboratory polysomnography for the management of more symptomatic patients with obstructive sleep apnea, but its effectiveness has not been evaluated across a broad clinical spectrum. OBJECTIVES: To compare the long-term effectiveness (6 mo) of home respiratory polygraphy and polysomnography management protocols in patients with intermediate-to-high sleep apnea suspicion (most patients requiring a sleep study). METHODS: A multicentric, noninferiority, randomized controlled trial with two open parallel arms and a cost-effectiveness analysis was performed in 12 tertiary hospitals in Spain. Sequentially screened patients with sleep apnea suspicion were randomized to respiratory polygraphy or polysomnography protocols. Moreover, both arms received standardized therapeutic decision-making, continuous positive airway pressure (CPAP) treatment or a healthy habit assessment, auto-CPAP titration (for CPAP indication), health-related quality-of-life questionnaires, 24-hour blood pressure monitoring, and polysomnography at the end of follow-up. The main outcome was the Epworth Sleepiness Scale measurement. The noninferiority criterion was -2 points on the Epworth scale. MEASUREMENTS AND MAIN RESULTS: In total, 430 patients were randomized. The respiratory polygraphy protocol was noninferior to the polysomnography protocol based on the Epworth scale. Quality of life, blood pressure, and polysomnography were similar between protocols. Respiratory polygraphy was the most cost-effective protocol, with a lower per-patient cost of 416.7€. CONCLUSIONS: Home respiratory polygraphy management is similarly effective to polysomnography, with a substantially lower cost. Therefore, polysomnography is not necessary for most patients with suspected sleep apnea. This finding could change established clinical practice, with a clear economic benefit. Clinical trial registered with www.clinicaltrials.gov (NCT 01752556).

Circulating blood levels of IL-6, IFN-γ, and IL-10 as potential diagnostic biomarkers in gastric cancer: a controlled study.

BACKGROUND: Gastric adenocarcinoma is the third most common cause of cancer-associated death worldwide. Helicobacter pylori infection activates a signaling cascade that induces production of cytokines and chemokines involved in the chronic inflammatory response that drives carcinogenesis. We evaluated circulating cytokines and chemokines as potential diagnostic biomarkers for gastric cancer. METHODS: We included 201 healthy controls and 162 patients with distal gastric cancer who underwent primary surgical resection between 2009 and 2012 in Mexico City. The clinical and pathological data of patients were recorded by questionnaire, and the cancer subtype was classified as intestinal or diffuse. Pathological staging of cancer was based on the tumor-node-metastasis staging system of the International Union Against Cancer. Concentrations of IL-1ß, IL-6, TNF-α, IL-10, and MCP-1 in serum were measured using multiplex analyte profiling technology and concentrations of IL-8, IFN-γ, and TGF-ß in plasma were measured using enzyme-linked immunosorbent assay. RESULTS: Levels of IL-1ß, IL-6, IFN-γ, and IL-10 were significantly higher and that of MCP-1 was lower in gastric cancer patients compared with controls. No differences in IL-8 or TNF-α levels were observed between gastric cancer and controls. IFN-γ and IL-10 were significantly higher in both intestinal and diffuse gastric cancer, whereas IL-1ß and IL-6 were higher and TGF-ß lower only in intestinal gastric cancer; MCP-1 was lower only in diffuse gastric cancer. IFN-γ and IL-10 levels were significantly higher in early (I/II) and late stage (III/IV) gastric cancer; IL-1ß and IL-8 were higher and MCP-1 was lower only in late stage (IV) patients. Receiver-operating characteristic analysis showed that for diagnosis of GC, IL-6 had high specificity (0.97) and low sensitivity (0.39), IL-10 had moderate specificity (0.82) and low sensitivity (0.48), and IL-1ß and IFN-γ showed low specificity (0.43 and 0.53, respectively) and moderate sensitivity (0.76 and 0.71, respectively). CONCLUSIONS: Increased levels of IL-6, IFN-γ, and IL-10 might be useful as diagnostic biomarkers for GC; however, this needs to be confirmed with larger number of patients and with control groups other than blood donors, properly age paired. IL-1ß, IL-6, MCP-1, and TGF-ß differentiate intestinal from diffuse GC. IFN-γ and IL-10 might be useful for diagnosis of early stage GC, and IL-1ß, IL-8, and MCP-1 for late stages of the disease.

Continuous Positive Airway Pressure Improves Quality of Life in Women with Obstructive Sleep Apnea. A Randomized Controlled Trial.

RATIONALE: Continuous positive airway pressure (CPAP) is the treatment of choice in patients with symptomatic obstructive sleep apnea (OSA). CPAP treatment improves quality of life (QoL) in men with OSA, but its role in women has not yet been assessed. OBJECTIVES: To investigate the effect of CPAP on QoL in women with moderate to severe OSA. METHODS: We conducted a multicenter, open-label randomized controlled trial in 307 consecutive women diagnosed with moderate to severe OSA (apnea-hypopnea index, ≥15) in 19 Spanish sleep units. Women were randomized to receive effective CPAP therapy (n = 151) or conservative treatment (n = 156) for 3 months. The primary endpoint was the change in QoL based on the Quebec Sleep Questionnaire. Secondary endpoints included changes in daytime sleepiness, mood state, anxiety, and depression. Data were analyzed on an intention-to-treat basis with adjustment for baseline values and other relevant clinical variables. MEASUREMENTS AND MAIN RESULTS: The women in the study had a mean (SD) age of 57.1 (10.1) years and a mean (SD) Epworth Sleepiness Scale score of 9.8 (4.4), and 77.5% were postmenopausal. Compared with the control group, the CPAP group achieved a significantly greater improvement in all QoL domains of the Quebec Sleep Questionnaire (adjusted treatment effect between 0.53 and 1.33; P < 0.001 for all domains), daytime sleepiness (-2.92; P < 0.001), mood state (-4.24; P = 0.012), anxiety (-0.89; P = 0.014), depression (-0.85; P = 0.016), and the physical component summary of the 12-item Short Form Health Survey (2.78; P = 0.003). CONCLUSIONS: In women with moderate or severe OSA, 3 months of CPAP therapy improved QoL, mood state, anxiety and depressive symptoms, and daytime sleepiness compared with conservative treatment. Clinical trial registered with www.clinicaltrials.gov (NCT02047071).

Modulation in the expression of SHP-1, SHP-2 and PTP1B due to the inhibition of MAPKs, cAMP and neutrophils early on in the development of cerulein-induced acute pancreatitis in rats.

The protein tyrosine phosphatases (PTPs) SHP-1, SHP-2 and PTP1B are overexpressed early on during the development of cerulein -induced acute pancreatitis (AP) in rats, and their levels can be modulated by some species of mitogen-activated protein kinases (MAPKs), the intracellular levels of cAMP and by general leukocyte infiltration, the latter at least for SHP-2 and PTP1B. In this study we show that cerulein treatment activates extracellular signal-regulated kinase (ERK) and c-Jun NH2-terminal kinase (JNK) but not p38 MAPK during the early phase of cerulein-induced AP (2h after the first injection of cerulein). Therefore, by using the MAPK inhibitors SP600125 (a specific JNK inhibitor) and PD98059 (a specific ERK inhibitor), we have unmasked the particular MAPK that underlies the modulation of the expression levels of these PTPs. JNK would act by preventing SHP-1 protein expression from increasing beyond a certain level. ERK 1/2 was the main MAPK involved in the increase in SHP-2 protein expression due to cerulein. JNK negatively modulated the SH2-domain containing PTPs. Both MAPKs played a role in the increase in PTP1B protein expression due to cerulein. Finally, by using the white blood cell inhibitors vinblastine sulfate, gadolinium chloride and FK506 (tacrolimus), we show that the macrophage activity or T-lymphocytes does not modulate the expression of any of the PTPs, although neutrophil infiltration was found to be a regulator of SHP-2 and PTP1B protein expression due to cerulein.

Tumoral EPAS1 (HIF2A) mutations explain sporadic pheochromocytoma and paraganglioma in the absence of erythrocytosis.

Pheochromocytomas (PCCs) and paragangliomas (PGLs) are chromaffin-cell tumors that arise from the adrenal medulla and extra-adrenal paraganglia, respectively. The dysfunction of genes involved in the cellular response to hypoxia, such as VHL, EGL nine homolog 1, and the succinate dehydrogenase (SDH) genes, leads to a direct abrogation of hypoxia inducible factor (HIF) degradation, resulting in a pseudo-hypoxic state implicated in PCC/PGL development. Recently, somatic post-zygotic mutations in EPAS1 (HIF2A) have been found in patients with multiple PGLs and congenital erythrocytosis. We assessed 41 PCCs/PGLs for mutations in EPAS1 and herein describe the clinical, molecular and genetic characteristics of the 7 patients found to carry somatic EPAS1 mutations; 4 presented with multiple PGLs (3 of them also had congenital erythrocytosis), whereas 3 were single sporadic PCC/PGL cases. Gene expression analysis of EPAS1-mutated tumors revealed similar mRNA EPAS1 levels to those found in SDH-gene- and VHL-mutated cases and a significant up-regulation of two hypoxia-induced genes (PCSK6 and GNA14). Interestingly, single nucleotide polymorphism array analysis revealed an exclusive gain of chromosome 2p in three EPAS1-mutated tumors. Furthermore, multiplex-PCR screening for small rearrangements detected a specific EPAS1 gain in another EPAS1-mutated tumor and in three non-EPAS1-mutated cases. The finding that EPAS1 is involved in the sporadic presentation of the disease not only increases the percentage of PCCs/PGLs with known driver mutations, but also highlights the relevance of studying other hypoxia-related genes in apparently sporadic tumors. Finally, the detection of a specific copy number alteration affecting chromosome 2p in EPAS1-mutated tumors may guide the genetic diagnosis of patients with this disease.

Pegylated interferon plus ribavirin in HIV-infected patients with recurrent hepatitis C after liver transplantation: a prospective cohort study.

BACKGROUND & AIMS: The aim of this study was to evaluate the results of treatment with pegylated interferon and ribavirin for the recurrence of hepatitis C after liver transplantation in HCV/HIV-coinfected patients. METHODS: This was a prospective, multicentre cohort study, including 78 HCV/HIV-coinfected liver transplant patients who received treatment for recurrent hepatitis C. For comparison, we included 176 matched HCV-monoinfected patients who underwent liver transplantation during the same period of time at the same centres and were treated for recurrent hepatitis C. RESULTS: Antiviral therapy was discontinued prematurely in 56% and 39% (p = 0.016), mainly because of toxicity (22% and 11%, respectively; p=0.034). Sustained virological response (SVR) was achieved in 21% of the coinfected patients and in 36% of monoinfected patients (p = 0.013). For genotype 1, SVR rates were 10% and 33% (p = 0.002), respectively; no significant differences were observed for the other genotypes. A multivariate analysis based on the whole series identified HIV-coinfection as an independent predictor of lack of SVR (OR, 0.17; 95% CI, 0.06-0.42). Other predictors of SVR were donor age, pretreatment HCV viral load, HCV genotype, and early virological response. SVR was associated with a significant improvement in survival: 5-year survival after antiviral treatment was 79% for HCV/HIV-coinfected patients with SVR vs. 43% for those without (p = 0.02) and 92% vs. 60% in HCV-monoinfected patients (p < 0.001), respectively. CONCLUSIONS: The response to pegylated interferon and ribavirin was poorer in HCV/HIV-coinfected liver recipients, particularly those with genotype 1. However, when SVR was achieved, survival of coinfected patients increased significantly.

Improved genetic counseling in Alport syndrome by new variants of COL4A5 gene.

There are current requirements of using genetic databases for offering a better genetic assistance to patients of some syndromes, especially those with X-linked heredity patterns (like Alport Syndrome) for the high probability of having descendants affected by the disease. We describe the first reported case of COL4A5 gene missense c.1499 G>T mutation in a 16-year-old girl confirmed to be affected by Alport Syndrome after genetic counseling. Next Generation Sequencing procedures let discover this mutation and offer an accurate clinical treatment to this patient. Current scientific understanding of genetic syndromes suggests the high importance of updated databases and the inclusion of Variant of Unknown Significance related to clinical cases. All of this updating could enable patients to have a better opportunity of diagnosis and having genetic and clinical counseling. This event is even more important in women planning to start a family to have correct genetic counseling regarding the risk posed to offspring, and allowing the decision to undergo prenatal testing.

Viral load, CMV-specific T-cell immune response and cytomegalovirus disease in solid organ transplant recipients at higher risk for cytomegalovirus infection during preemptive therapy.

Despite advances in prevention, cytomegalovirus (CMV) recurrence is an important challenge in high-risk organ recipients. The present study prospectively evaluates the impact of CMV-specific T-cell immune response and secondary prophylaxis on the risk of recurrence in a cohort of CMV high-risk organ recipients and whether it is possible to determine a safe standardized viral load value below which CMV disease is unlikely. Thirty-nine recipients were included. Thirty-six had primary infections, and 88.9% recurred. Rate and duration of recurrent CMV infection was similar in patients with and without secondary prophylaxis: 57.9% vs. 53.6%, P = 0.770 and 16 vs. 15 days, P = 0.786, respectively. The only factor independently associated with no episodes of CMV recurrence was the acquisition of CMV-specific T-cell immune response (OR: 0.151, 95% CI: 0.028-0.815; P = 0.028). Cytomegalovirus diseases (N = 5) occurred in patients with CMV viral load above 1500 IU/ml who did not follow the planned monitorization schedule. Our observations suggest that episodes of recurrent CMV infection are common after preemptive therapy despite secondary prophylaxis and that CMV-specific T-cell immune response is associated with a decreased risk of recurrent infections. Preemptive therapy may be safe in patients at high risk for CMV infection with strict close monitoring of the CMV viral load.

High expression of Toll-like receptors 2 and 9 and Th1/Th2 cytokines profile in obese asthmatic children.

Asthma is a common pulmonary disease with chronic inflammation of the airways, and obesity is a chronic state of low-grade inflammation. Toll-like receptors (TLRs) are involved in the innate immune response. This study was designed to analyze whether obesity has an effect on the immune response of patients with asthma. We included obese asthmatic, obese, asthmatic, and healthy children. Biochemical and anthropometric analyses were performed. Interleukin (IL)-2, interferon (IFN) gamma, IL-4, IL-10, IL-1beta, and tumor necrosis factor alpha were measured. Peripheral blood mononuclear cells were analyzed by immunostaining with anti-TLR2 and anti-TLR9 antibodies. The data were expressed as means ± SEM or medians and percentiles. Kruskal-Wallis test and Dunn's multiple comparison test were applied. Asthmatic patients, both obese and nonobese, exhibited a mild asthma phenotype; none had infectious process, exacerbation, or acute symptoms during the 30 days before the inclusion in the study. The IL-2 and IFN-gamma levels in the obese asthmatic group were lower than in the other three groups. IL-4 levels in the obese asthmatic group were almost equal to those of the asthmatic group and more than in the other two groups, without significant difference. There were higher levels of TLR2 and TLR9 in obese asthmatic patients than in the other three groups. There is a decrease in Th1 cytokines in obese asthmatic patients, and we only found a trend to an increased Th2 profile. Patients studied do not appear to fit into any of the endotypes described until now. This is the first study showing the high expression of TLR2 and TLR9 in obese asthmatic patients. It is necessary to study other cytokines in obese asthmatic patients to see if it is possible to fit them into any of the already described endotypes or if it is a distinct endotype.

Proteomic analysis of the soluble and the lysosomal+mitochondrial fractions from rat pancreas: Implications for cerulein-induced acute pancreatitis.

Alterations in protein expression within the initiation phase of acute pancreatitis (AP) might play an important role in the development of this disease, lysosomes being involved in its pathophysiology. The use of pancreatic subcellular fractions in proteomic analysis, simplifies protein maps and helps in the identification of new protein changes and biomarkers characterizing tissue damage. The present study aims to determine the differentially expressed acidic proteins in the pancreatic soluble and lysosomal+mitochondrial (L+M) fractions from rats during the early phase of the experimental model of cerulein (Cer)-induced AP. Subcellular pancreatic extracts from diseased and control rats were analyzed by 2-DE (3-5.6 pH range) and MALDI-TOF/TOF MS. Comparative analysis afforded the conclusive identification of 13 (soluble fraction) and 7 (L+M fraction) proteins or protein fragments occuring in different amounts between diseased and control pancreas, some of them being newly described in AP. In the soluble fraction, we detected changes related to inflammation and apoptosis (α1-inhibitor-3, α-1 antitrypsin, α-1 macroglobulin, haptoglobin, STRAP), oxidative stress and stress response (peroxiredoxin-2, thioredoxin-like 1, GRP94/TRA1, heat shock cognate 71kDa protein), digestive proteases (elastase 3B), serine protease inhibition (serpins B6 and A3L) and translation processes (EF 1-δ). In the L+M fraction, we detected changes mainly related to energy generation or cellular metabolism (ATP synthase ß subunit, chymotrypsinogen B, triacylglycerol lipase), cell redox homeostasis (iodothyronine 5´monodeiodinase) and digestive proteases (carboxypeptidase B1). The data should provide valuable information for unraveling the early pathophysiologic mechanisms of Cer-induced AP.

Toll-like receptors and cytokines are upregulated during Helicobacter pylori infection in children.

BACKGROUND AND AIM: Helicobacter pylori infection is mainly acquired during childhood, and establishes a chronic infection that may lead to peptic ulcer or gastric cancer during adulthood. Toll-like receptors (TLRs) are expressed by distinct cell types throughout the gastrointestinal tract, and play an important role in regulation of the innate immune response. Few works have addressed TLRs expression in gastric epithelia of adults, and scarce studies have done it in children. The aim of this work was to analyze the expression of TLR2, TLR4, TLR5, TLR9, and IL-8, IL-10 and TNF-α in the gastric mucosa of children with and without H. pylori infection. METHODS: Gastric biopsies were collected by endoscopy from 50 children with recurrent abdominal pain, 25 with H. pylori infection and 25 without infection. In the gastric biopsies the expression of TLRs and cytokines was studied by immunohistochemistry, and the degree of mucosal inflammation was determined using the Sydney system. RESULTS: We found that H. pylori infection was associated with a significant increased expression of TLRs 2, 4, 5 and 9, although expression varied between surface epithelia and glands. Epithelial cells expressing IL-8, IL-10 and TNF-α were increased in gastric mucosa of children with H. pylori infection. CONCLUSION: This study shows the gastric epithelia of children respond to H. pylori infection by increasing the expression of TLR2, TLR4, TLR5, TLR9 and the cytokines IL-8, IL-10 and TNF-α.