Abscisic Acid-Triggered Persulfidation of the Cys Protease ATG4 Mediates Regulation of Autophagy by Sulfide.

Hydrogen sulfide is a signaling molecule that regulates essential processes in plants, such as autophagy. In Arabidopsis (Arabidopsis thaliana), hydrogen sulfide negatively regulates autophagy independently of reactive oxygen species via an unknown mechanism. Comparative and quantitative proteomic analysis was used to detect abscisic acid-triggered persulfidation that reveals a main role in the control of autophagy mediated by the autophagy-related (ATG) Cys protease AtATG4a. This protease undergoes specific persulfidation of Cys170 that is a part of the characteristic catalytic Cys-His-Asp triad of Cys proteases. Regulation of the ATG4 activity by persulfidation was tested in a heterologous assay using the Chlamydomonas reinhardtii CrATG8 protein as a substrate. Sulfide significantly and reversibly inactivates AtATG4a. The biological significance of the reversible inhibition of the ATG4 by sulfide is supported by the results obtained in Arabidopsis leaves under basal and autophagy-activating conditions. A significant increase in the overall ATG4 proteolytic activity in Arabidopsis was detected under nitrogen starvation and osmotic stress and can be inhibited by sulfide. Therefore, the data strongly suggest that the negative regulation of autophagy by sulfide is mediated by specific persulfidation of the ATG4 protease.

Covid-19 transmission, outcome and associated risk factors in cancer patients at the first month of the pandemic in a Spanish hospital in Madrid.

BACKGROUND: There are no large reported series determining the Covid-19 cancer patient's characteristics. We determine whether differences exist in cumulative incidence and mortality of Covid-19 infection between cancer patients and general population in Madrid. MATERIAL AND METHODS: We reviewed 1069 medical records of all cancer patients admitted at Oncology department between Feb 1 and April 7, 2020. We described Covid-19 cumulative incidence, treatment outcome, mortality, and associated risk factors. RESULTS: We detected 45/1069 Covid-19 diagnoses in cancer patients vs 42,450/6,662,000 in total population (p < 0.00001). Mortality rate: 19/45 cancer patients vs 5586/42,450 (p = 0.0001). Mortality was associated with older median age, adjusted by staging and histology (74 vs 63.5 years old, OR 1.06, p = 0.03). Patients who combined hydroxychloroquine and azithromycin presented 3/18 deaths, regardless of age, staging, histology, cancer treatment and comorbidities (OR 0.02, p = 0.03). CONCLUSION: Cancer patients are vulnerable to Covid-19 with an increase in complications. Combined hydroxychloroquine and azithromycin is presented as a good treatment option.

Fast update of undetected Mycobacterium chimaera infections to reveal unsuspected cases.

Mycobacterium chimaera is involved in a worldwide alert due to contaminated heater-cooler units. A real-time polymerase chain reaction (RT-PCR)-based procedure was implemented to survey undetected cases of M. chimaera infection. PCR was negative in the 59 prosthetic heart valves from patients with PCR-16SrRNA-negative infective endocarditis. PCR identified M. chimaera in one of 15 clinically significant retrospective Mycobacterium avium-Mycobacterium intracellulare complex isolates, which corresponded to a patient who had undergone heart valve replacement in a different institution. Whole-genome sequencing demonstrated that he was the first case in Spain with involvement of the strain responsible for the global outbreak. These results highlight the relevance of retrospective tracking for undetected M. chimaera infections.

Impact of interferon-free regimens on the glomerular filtration rate during treatment of chronic hepatitis C in a real-life cohort.

Little data are available on renal toxicity exerted by direct-acting antivirals (DAAs) in real life. The aim of this study was to assess the impact of direct-acting antivirals against hepatitis C virus infection currently used in Spain and Portugal on the estimated glomerular filtration rate (eGFR) in clinical practise. From an international, prospective multicohort study, patients treated with DAAs for at least 12 weeks and with eGFR ≥30 mL/min per 1.73 m2 at baseline were selected. eGFR was determined using the CKD-EPI formula. A total of 1131 patients were included; 658 (58%) were HIV/HCV-coinfected patients. Among the 901 patients treated for 12 weeks, median (interquartile range) eGFR was 100 (87-107) at baseline vs 97 (85-105) mL/min per 1.73 m2 at week 12 of follow-up (FU12) post-treatment (P < .001). For HIV-coinfected subjects who received tenofovir plus a ritonavir-boosted HIV protease inhibitor (PI/r), baseline vs FU12 eGFR were 104 (86-109) vs 104 (91-110) mL/min per 1.73 m2 (P = .913). Among subjects receiving ombitasvir/paritaprevir with or without dasabuvir, eGFR did not show any significant change. Of 1100 subjects with eGFR >60 mL/min per 1.73 m2 at baseline, 22 (2%) had eGFR <60 mL/min per 1.73 m2 at FU12, but none presented with eGFR <30 mL/min per 1.73 m2 . In conclusion, eGFR slightly declines during therapy with all-oral DAAs and this effect persists up to 12 weeks after stopping treatment in subjects with normal to moderately impaired renal function, regardless of HIV status. Concomitant use of tenofovir plus PI/r does not seem to have an impact on eGFR.

Identification of aspartic acid-203 in human thymidine phosphorylase as an important residue for both catalysis and non-competitive inhibition by the small molecule "crystallization chaperone" 5'-O-tritylinosine (KIN59).

Thymidine phosphorylase (TP) is a catabolic enzyme in thymidine metabolism that is frequently upregulated in many solid tumors. Elevated TP levels are associated with tumor angiogenesis, metastasis and poor prognosis. Therefore, the use of TP inhibitors might offer a promising strategy for cancer treatment. The tritylated inosine derivative 5'-O-tritylinosine (previously designated KIN59) is a non-competitive inhibitor of TP which was previously found to be instrumental for the crystallization of human TP. A combination of computational studies including normal mode analysis, automated ligand docking and molecular dynamics simulations were performed to define a plausible binding site for 5'-O-tritylinosine on human TP. A cavity in which 5'-O-tritylinosine could fit was identified in the vicinity of the Gly405-Val419 loop at a distance of about 11A from the substrate-binding site. In the X-ray crystal structure, this pocket is characterized by an intricate hydrogen-bonding network in which Asp203 was found to play an important role to afford the loop stabilization that is required for efficient enzyme catalysis. Site-directed mutagenesis of this amino acid residue afforded a mutant enzyme with a severely compromised catalytic efficiency (V(max)/K(m) of mutant enzyme approximately 50-fold lower than for wild-type TP) and pronounced resistance to the inhibitory effect of 5'-O-tritylinosine. In contrast, the D203A mutant enzyme kept full sensitivity to the competitive inhibitors 6-aminothymine and 6-amino-5-bromouracil, which is in line with the kinetic properties of these inhibitors. Our findings reveal the existence of a previously unrecognized site in TP that can be targeted by small molecules to inhibit the catalytic activity of TP.

Thymidine phosphorylase is noncompetitively inhibited by 5'-O-trityl-inosine (KIN59) and related compounds.

We found that 5'-O-trityl-inosine (KIN59) inhibits recombinant bacterial (E. coli) and human thymidine phosphorylase (TPase) with an IC50 of 44 microM and 67 microM, respectively. In contrast to previously described TPase inhibitors, KIN59 does not compete with thymidine (dThd) at the pyrimidine nucleoside-binding site or with inorganic phosphate (Pi) at the phosphate-binding site of the enzyme. These findings are strongly suggestive for the presence of an allosteric binding site at the enzyme. TPase is identical to the angiogenic protein platelet-derived endothelial cell growth factor (PD-ECGF). As such, PD-ECGF stimulates angiogenesis in the chick chorioallantoic membrane (CAM) assay. This angiogenic response was completely inhibited by KIN59. Inosine did not inhibit the enzyme or the angiogenic effect of TPase, confirming that the 5'-O-trityl group in KIN59 is essential for the observed effect. Our observations indicate that allosteric sites in TPase may regulate its biological activity.

Expression of the neurotrophin receptor TrkB in the mouse liver.

Neurotrophins acting through Trk signal-transducing receptors play essential roles in the nervous system, and probably in some non-neuronal tissues. In the present study, we used RT-PCR, Western-blot and immunohistochemistry to investigate the occurrence and cellular localization of TrkB in the mouse liver, from newborns to 6 months. Furthermore, the structure of the liver in mice carrying a mutation in the trkB gene, resulting in a non-functional protein, was studied. The analysis of the DNA sequence showed that hepatic trkB gene is identical to the cerebral one, and TrkB mRNA and TrkB full-length protein (145 kDa) were detected at all the ages sampled. Immunohistochemistry revealed age-dependent changes in the pattern of TrkB expression. From 0 to 15 days, the TrkB was detected in morphologically and immunohistochemically identified monocyte-macrophage-dendric cells scattered throughout the organ, while in animals 3- and 6-months-old it was restricted to nerve fibres. Interestingly, there was a parallelism between TrkB expression by monocyte-macrophage-dendric cells and the presence of hepatic erythroblastic islands. In agreement with a possible role of TrkB on hepatic haematopoiesis, the liver of 15 days old TrkB (-/-) mice still contained erythroblastic islands, whereas they were absent in the wild-type littermates. Another striking finding was the absence of nerve profiles in the TrkB (-/-) animals. All together, present results support the role of TrkB in the murine liver in maintaining the innervation of the organ, and more importantly throughout an unknown mechanism in controlling the hepatic haematopoietic function.

[Xanthogranulomatous pyelonephritis in childhood. A case report]. / Pielonefritis xantogranulomatosa en la infancia. A propósito de un caso.

Xanthogranulomatous pyelonephritis is an uncommon chronic inflammatory renal disorder. Most cases have been described in middle aged women and it is extremely rare in children. We report a case of a 4 year old girl who suffered from recurrent urinary tract infection and antibiotic therapy resistance. She had low growth-rate and palpable left renal mass on examination and was confirmed by radiological findings. We performed left kidney partial resection and then histological examination showed focal xanthogranulomatous pyelonephritis. The focal form of the disease may respond to antibiotic treatment although usually an enucleation or partial resection must be performed. In conclusion xanthogranulomatous pyelonephritis should be considered in the differential diagnosis of a renal mass and recurrent urinary tract infection in childhood.

[Kidney pelvis hemangioma as a cause of hematuria]. / Angiomatosis pélvica como causa de hematuria.

UNLABELLED: Kidney hemangiomas are congenital vascular anomalies with an unknown ethiology. They are considered as benign tumours with a poor clinic expression (hematuria or ureteric colic, which is associated to the drop of clots in the urinary tract). OBJECTIVE: To expound a case of unilateral severe hematuria in a young woman with a restless past medical history, due to a pelvic left kidney hemangioma. MATERIALS AND METHODS: Clinic case of a sixteen years old woman, no relevant past medical history, presented with an anaemizing gross hematuria and passing vermiform clots. As diagnostic techniques we used ultrasonography, excretory urography, cystourethroscopy, CT, angiography and nuclear imaging. Surgery (flank nephectomy) was needed for treatment. RESULTS: Pathology report: submucosal vascular proliferative process in kidney pelvis composed by blood vessels, suggesting haemangioma.

TrkB mRNA and protein in mouse spleen: structure of the spleen of functionally deficient TrkB mice.

Whereas it is nowadays clear that neurotrophins are involved in the regulation of various aspects of the functioning of immune system, knowledge of their actual immunomodulatory roles is still fragmentary and incomplete. In this respect, knock-out mouse models remain particularly unexplored. In the present study, the expression of the TrkB neurotrophin receptor in murine spleen was addressed at the mRNA (reverse transcription/polymerase chain reaction) and protein (Western blot) levels. Once the presence of TrkB at both levels was demonstrated, the age-dependent changes in the pattern of expression of the receptor were analyzed and quantified, and TrkB-positive cells were identified by immunohistochemistry. TrkB-immunoreactive cells, identified as red pulp macrophages, were detected in the spleen throughout postnatal development and adult life; their numbers peaked at the age of 15 days. The absence of functional TrkB did not appear to result in morphological changes as assessed by light and electron microscopy of spleens from 15-day-old mice knockout for the trkB gene. The present results support the idea that, in the murine spleen, TrkB and its ligands are involved in macrophage physiology in a developmentally regulated fashion, but they do not seem to be essential for macrophage survival.

[Fibroepithelial polyp of the urethra in a young woman]. / Pólipo fibroepitelial de uretra en mujer joven.

OBJECTIVE: We present the case of a fibroepithelial polyp from the uretra of a young female, analysing the factors that make this an interesting case. MATERIAL AND METHODS: A case-report of a fibrous polyp from the urethra of a young female, an analysis of its differential diagnosis and a review of the literature. RESULTS AND CONCLUSIONS: The fibrous polyp of the urethra (fibroepithelual polyp, simple polyp, pedunculated polyp) is a rare benign epithelial tumour, more frequently encountered in males and in the posterior urethra. When it appears in females it is usually located in the external meatus and normally asymptomatic, although it can also present with a mictional syndrome, terminal hematuria or urethral hemorrhage. The differential diagnosis includes other benign epithelial tumours of the urethra, such as the caruncula, mainly the granulomatous, very difficult to differentiate, the squamous papilloma and the acuminated chondiloma. A clinical examination and histological study are necessary for a correct diagnosis, and treatment consists of surgical resection.

[Value of post-miction residue after urethrocervicopexy. Prospective study]. / Valor del residuo posmiccional tras la uretrocervicopexia. Estudio prospectivo.

OBJECTIVE: To demonstrate that a high post-micturitional residue after a colposuspension is not a success predictive factor in the incontinence surgery. METHODS: We designed two identical groups of work, with the only difference of a high post-micturitional residue in one of them after the surgery (< 100 cc), and we evaluated the surgical results, with a follow-up of one year. We considered fails the presence of losses related with stress. RESULTS: There were no significative differences in the number of the deliveries, age, weight, length or body mass index in both groups. There were significative differences in the permanence of the urethral catheter and residue of urine, p < 0.001. There were no differences in the number of success in both groups. CONCLUSION: A high post-micturitional residue after colposuspension is not an influent factor in the final result of surgery.

[Urethrocervicopexy in aged women. Prospective study]. / Uretrocervicopexia en la mujer mayor. Un estudio prospectivo.

OBJECTIVES: [corrected] To demonstrate that Burch colposuspension gives a higher index of fails in over sixty years women affected of stress incontinence than in women under sixty years of age. METHODS: We established two groups of follow-up, each of them included twenty women affected of stress urine incontinence and surgical treatment indication. The only difference between both groups was the age of the patients, with a limit established in sixty years. We have evaluated the results of surgical intervention one year after it. The binomial parameters are compared with x2 test; the continuous parameters with the t-Student test for independent parameters. RESULTS: We did not find statistical significative differences with number of deliveries, body mass index, previous pelvic surgery, perineal rehabilitation, or the importance of the incontinence, between both groups of women. We found statistical significative difference in the novo micturition urgency or an increase in a previous urgency (p < 0.1) in women over sixty years. We did not find statistical significative differences in the number of successes and fails in both groups. CONCLUSION: The age is not a risk factor of failing of Burch colposuspension, but it is a risk factor of the appearance or increase of micturition urgency.

Distribución, abundancia, densidad y morfometría de Strombus gigas (Mesogasteropoda: Strombidae) en el Arrecife Alacranes, Yucatán, México / Distribution, abundance, density and morphometry of Strombus gigas (Mesogasteropoda: Strombidae) in Alacranes reef, Yucatán, México

A population of the conch Strombus gigas was sampled in 109 transects (100 x 10 m each) for an area of 109000 m2 (eight surveys in 1998 and six in 1999) in Yucatan, Mexico; S. gigas was the most abundant conch species (97% of individuals) and ocurred at depths of 0.5-18 m. Mean density was 0.0084 individuals/m2 (Standard deviation SD=0.0138, n=1910). Maximum density (0.015 individuals/m2) was at 0.5-2 m and minimal (0.001 individuals/m2) at 16-18 m. Shell length was 120-290 mm (mena=211.6, SD=26, n=1136). Shell lip thickness, measured only in adults, was 1-28 mm (mean=8.7, SD=7.5, n=1136).

Novel ribofuranosylnucleoside lead compounds for potent and selective inhibitors of mitochondrial thymidine kinase-2.

The ribonucleoside analogues (E)-5-(2-bromovinyl)uridine (5-BV-Urd) and 3'-spiro-(4'-amino-1',2'-oxathiole-2',2'-dioxide)-5-methyluridine (3'-AOD-5-MeUrd) emerged as potent and selective competitive inhibitors of mitochondrial thymidine kinase (TK)-2 with respect to thymidine (K(i)/K(m) values of 9.0 and 1.2 respectively). Cytosolic TK-1 did not show measurable affinity for these compounds. [(32)P]Phosphate transfer studies from [gamma-(32)P]ATP to 5-BV-Urd and 3'-AOD-5-MeUrd revealed extremely poor substrate activity but potent inhibitory potential of the compounds. It was concluded that the ribonucleosides 5-BV-Urd and 3'-AOD-5-MeUrd represent two new lead compounds for potent and selective inhibitors of mitochondrial TK-2.

Changes in the expression of the nerve growth factor receptors TrkA and p75LNGR in the rat thymus with ageing and increased nerve growth factor plasma levels.

The nerve growth factor (NGF) receptors p75LNGR and TrkA are expressed by thymic epithelial cells. Presumably, the NGF-TrkA system is involved in the paracrine communication between thymic epithelial cells and thymocytes, whereas the functional role of p75LNGR is still unknown. The thymus of vertebrates undergoes age-related changes that in part depend on hormonal factors. In order to find out whether thymic epithelial cells are responsive to NGF during the whole lifespan of the rat, we studied NGF receptor expression in the thymus from birth to 2 years of age, using immunohistochemistry. Furthermore, to evaluate whether increased plasma levels of NGF affected the ageing process, either NGF or 4-methylcatechol (4MC), an inductor of NGF synthesis, was administered. Both TrkA and p75LNGR were expressed by a subpopulation of thymic epithelial cells during the whole age range studied and their expression peaked at around 3 months. TrkA was primarily found in subcortical and medullary epithelial cells, whereas p75LNGR was seen in a subpopulation of medullary cells. Cortical epithelial cells, neural crest-derived cells, other stromal cells and thymocytes were not immunoreactive for NGF receptors. Neither the administration of NGF nor the increased NGF plasma levels obtained after 4MC treatment seemed to affect the ageing of the thymus as assessed by morphological and immunohistochemical criteria, but this increase in NGF levels did produce a shift in the expression of p75LNGR from epithelial cells to ED1-positive macrophages in animals of 6 months and older. Present results indicate that the expression of p75LNGR and TrkA in the rat thymus undergoes age-dependent changes that parallel those of epithelial cells. NGF could therefore be important for thymus homeostasis, possibly acting on epithelial cells. Nevertheless, NGF did not seem to be able to prevent the involution of this organ, although it produced a switch in the expression of p75LNGR, the significance of which remains to be established.

[Nephrogenic adenoma. Report of 4 new cases]. / Adenoma nefrogénico. Presentación de cuatro nuevos casos.

OBJECTIVE: Four additional cases of nephrogenic adenoma are described. METHODS: The clinical records of patients with nephrogenic adenoma that had been diagnosed at our hospital from 1994 to 1999 were reviewed. RESULTS: The most frequent localization was the urinary bladder and it was more prevalent in middle-aged men. This condition usually presented as a histological finding in the course of follow-up or treatment of other bladder conditions by TUR or cystectomy. Biopsy is fundamental for diagnosis. CONCLUSIONS: Nephrogenic adenoma is a rare benign metaplastic lesion of the urinary tract that appears to result from chronic irritative stimulus to the urothelium. The treatment of choice is by transurethral resection. Although no case of progression to malignancy have been detected, long-term follow-up by endoscopy and ultrasound is advocated due to its predispositon to recurrence.

Expression of the neurotrophin receptor TrkB in rat spleen macrophages.

Increasing evidence suggests that some members of the family of the neurotrophins could be involved in immune system functioning. Both neurotrophins and their tyrosine-kinase signal-transducing receptors, the so-called Trk receptors, have been detected in various lymphoid tissues in a number of species. Nevertheless, their cellular localisation remains unclear in most cases. In this study, we used immunohistochemical techniques to localise TrkB in the rat spleen (from 0 days to 2 years). Cells expressing TrkB-like immunoreactivity were found exclusively within the white pulp of the spleen, along the marginal zone-follicle border and inside the follicles and periarteriolar lymphoid sheaths. These cells probably represented macrophage subpopulations, since they expressed the ED3 rat macrophage antigen. No evidence of TrkB-like protein expression in lymphocytes or follicular dendritic cells could be found. Furthermore, the density of TrkB-immunoreactive cells was observed to increase with age. Although the role of TrkB ligands in these cells remains to be clarified, the present findings provide further evidence for the supposed role of neurotrophins in immune system homeostasis.