Unveiling mutation effects on the structural dynamics of the main protease from SARS-CoV-2 with hybrid simulation methods.

The main protease of SARS-CoV-2 (called Mpro or 3CLpro) is essential for processing polyproteins encoded by viral RNA. Several Mpro mutations were found in SARS-CoV-2 variants, which are related to higher transmissibility, pathogenicity, and resistance to neutralization antibodies. Macromolecules adopt several favored conformations in solution depending on their structure and shape, determining their dynamics and function. In this study, we used a hybrid simulation method to generate intermediate structures along the six lowest frequency normal modes and sample the conformational space and characterize the structural dynamics and global motions of WT SARS-CoV-2 Mpro and 48 mutations, including mutations found in P.1, B.1.1.7, B.1.351, B.1.525 and B.1.429+B.1.427 variants. We tried to contribute to the elucidation of the effects of mutation in the structural dynamics of SARS-CoV-2 Mpro. A machine learning analysis was performed following the investigation regarding the influence of the K90R, P99L, P108S, and N151D mutations on the dimeric interface assembling of the SARS-CoV-2 Mpro. The parameters allowed the selection of potential structurally stable dimers, which demonstrated that some single surface aa substitutions not located at the dimeric interface (K90R, P99L, P108S, and N151D) are able to induce significant quaternary changes. Furthermore, our results demonstrated, by a Quantum Mechanics method, the influence of SARS-CoV-2 Mpro mutations on the catalytic mechanism, confirming that only one of the chains of the WT and mutant SARS-CoV-2 Mpros are prone to cleave substrates. Finally, it was also possible to identify the aa residue F140 as an important factor related to the increasing enzymatic reactivity of a significant number of SARS-CoV-2 Mpro conformations generated by the normal modes-based simulations.

Triacontyl p-coumarate: an inhibitor of snake venom metalloproteinases.

Snake venom metalloproteinases (SVMPs) participate in a number of important biological, physiological and pathophysiological processes and are primarily responsible for the local tissue damage characteristic of viperid snake envenomations. The use of medicinal plant extracts as antidotes against animal venoms is an old practice, especially against snake envenomations. Such plants are sources of many pharmacologically active compounds and have been shown to antagonize the effects of some venoms and toxins. The present study explores the activity of triacontyl p-coumarate (PCT), an active compound isolated from root bark of Bombacopsis glabra vegetal extract (Bg), against harmful effects of Bothropoides pauloensis snake venom and isolated toxins (SVMPs or phospholipase A(2)). Before inhibition assays, Bg or PCT was incubated with venom or toxins at ratios of 1:1 and 1:5 (w/w; venom or isolated toxins/PCT) for 30 min at 37°C. Treatment conditions were also assayed to simulate snakebite with PCT inoculated at either the same venom or toxin site. PCT neutralized fibrinogenolytic activity and plasmatic fibrinogen depletion induced by B. pauloensis venom or isolated toxin. PCT also efficiently inhibited the hemorrhagic (3MDH - minimum hemorrhagic dose injected i.d into mice) and myotoxic activities induced by Jararhagin, a metalloproteinase from B. jararaca at 1:5 ratio (toxin: inhibitor, w/w) when it was previously incubated with PCT and injected into mice or when PCT was administered after toxin injection. Docking simulations using data on a metalloproteinase (Neuwiedase) structure suggest that the binding between the protein and the inhibitor occurs mainly in the active site region causing blockade of the enzymatic reaction by displacement of catalytic water. Steric hindrance may also play a role in the mechanism since the PCT hydrophobic tail was found to interact with the loop associated with substrate anchorage. Thus, PCT may provide a alternative to complement ophidian envenomation treatments.

Gait pattern classification in children with Charcot-Marie-Tooth disease type 1A.

Gait pattern classification may assist in clinical decision making and cluster analysis (CA) has been often adopted to this aim. The goal of this study was to identify, through CA, typical walking patterns in a group of 21 young subjects with CMT1A, a hereditary progressive neuropathy, and to study possible correlation with the disease's clinical status. The protocol included kinematic/kinetic analysis of natural walking and more demanding locomotor tasks, i.e. toe- and heel-walking. Hierarchical cluster analysis was carried out on parameters related to primary signs (foot-drop and push-off deficit) and, separately, to compensatory mechanisms at proximal (pelvis, hip and knee) or distal (ankle) level. CA on primary signs during natural walking identified three clusters: (1) pseudo-normal patients (PN), not significantly different from controls; (2) patients showing only foot-drop (FD); (3) patients with foot-drop and push-off deficit (FD&POD). Patients belonging to the PN subgroup showed distal abnormalities during heel-walking. The FD&POD subgroup was associated to a significantly worse clinical score (CMTES, p<0.05). The main compensatory strategies, which occurred independently from primary clusterization, included augmented hip/knee flexion in swing (steppage) and early ankle plantarflexion at mid stance (vaulting). We concluded that, although a number of young CMT1A patients do not show typical primary deviations during natural walking, they do show significant abnormalities in more demanding locomotor tasks that should be therefore considered. It is also hypothesized that progression of this degenerative condition may be associated to the migration of patients to more severe clusters, with possible appearance of compensatory strategies.

An unusual cause of chest pain in a young man: bronchogenic cysts and their cardiac manifestations.

We report a case of a 24-year-old man who presented with chest pain and electrocardiographic evidence of myocardial ischaemia. An abnormal structure located behind the heart on the urgent transthoracic echocardiogram and a computed tomography scan of the mediastinum led to prompt surgery with eventual resection of the lesion. The histology revealed fragments of connective tissue covered by squamous epithelium and ciliated epithelium, consistent with a bronchogenic cyst. The case study is accompanied by a literature review of the pathogenesis, diagnosis and management of bronchogenic cysts and their association with cardiac symptoms.

Phylogeny of ladybirds (Coleoptera: Coccinellidae): are the subfamilies monophyletic?

The Coccinellidae (ladybirds) is a highly speciose family of the Coleoptera. Ladybirds are well known because of their use as biocontrol agents, and are the subject of many ecological studies. However, little is known about phylogenetic relationships of the Coccinellidae, and a precise evolutionary framework is needed for the family. This paper provides the first phylogenetic reconstruction of the relationships within the Coccinellidae based on analysis of five genes: the 18S and 28S rRNA nuclear genes and the mitochondrial 12S, 16S rRNA and cytochrome oxidase subunit I (COI) genes. The phylogenetic relationships of 67 terminal taxa, representative of all the subfamilies of the Coccinellidae (61 species, 37 genera), and relevant outgroups, were reconstructed using multiple approaches, including Bayesian inference with partitioning strategies. The recovered phylogenies are congruent and show that the Coccinellinae is monophyletic but the Coccidulinae, Epilachninae, Scymninae and Chilocorinae are paraphyletic. The tribe Chilocorini is identified as the sister-group of the Coccinellinae for the first time.

Patients' perception of cancer-related fatigue: results of a survey to assess the impact on their everyday life

PURPOSE: Fatigue is a cancer-related symptom with great impact on patients' daily lives, but often not discussed with their oncologists. This survey explored functional and psychological fatigue impact among different cancer symptoms according to patient's perception (pp). METHODS: A cross-sectional, self-administered survey was conducted in 10 oncologist services throughout Spain. Demographical data and tumour diagnoses were collected. Fatigue impact on functional and social activities (Likert scale) and on emotional well-being (visual analogue scale) was measured. The pp of oncologist's response to fatigue report was recorded. RESULTS: 505 surveyed cancer patients were analysed (55.2% women, aged 58.8 years +/-11.7), 97.8% remembered experiencing fatigue during treatment. 27.1% did not discuss their fatigue with their oncologist. Fatigue affected patient's daily routine (> or = 50% of times) included self-care (58.26%), entertainment activities (69.8%), and relationships (71.4%). Fatigue was the most bothersome symptom of cancer. CONCLUSIONS: Cancer patients perceive fatigue as the symptom with highest impact on their daily living and that substantially affects their emotional and social areas (AU)

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Preliminary X-ray crystallographic studies of a tetrameric phospholipase A2 formed by two isoforms of crotoxin B from Crotalus durissus terrificus venom.

Crotoxin B is a basic phospholipase A2 found in the venom of Crotalus durissus terrificus and is one of the subunits that constitute crotoxin. This heterodimeric toxin, which is the main component of C. d. terrificus venom, is completed by an acidic, nontoxic and non-enzymatic component (crotoxin A) and is involved in important envenomation effects, such as neurological disorders, myotoxicity and renal failure. Although crotoxin was first crystallized in 1938, no crystal structure is currently available for crotoxin, crotoxin A or crotoxin B. In this work, the crystallization, X-ray diffraction data collection to 2.28 A resolution and molecular-replacement solution of a novel tetrameric complex formed by two dimers of crotoxin B isoforms (CB1 and CB2) is presented.

Down-regulation of vinculin upon MK886-induced apoptosis in LN18 glioblastoma cells.

Glioblastomas are a type of malignant brain tumor and are among the most difficult cancers to treat. One strategy to treat aggressive cancers is the use of drugs that target multiple signaling pathways. MK886 is a drug known to inhibit both 5- lipoxygenase-activating-protein (FLAP) and peroxisome proliferator activated receptor-alpha (PPAR-alpha). The objectives of this study were to investigate the ability of MK886 to induce apoptotic cell death in LN18 glioblastoma cells and to characterize the cell death mechanisms. MK886 induced massive apoptotic LN18 cell death that was manifested by the release of nucleosomes, annexinV binding to phosphatidylserine in the absence of nuclear staining, and changes in the fluorescent intensity of Mito Tracker Deep Red 633 indicating changes in mitochondrial oxidative function and mass. The alteration of the mitochondrial function implied that MK886 induced apoptosis in LN18 cells via a mitochondrial pathway. The broad caspases inhibitor ZVAD-FMK inhibited MK886-induced nucleosome release, but not annexinV binding or MK886-altered mitochondrial function. Real time RT-PCR demonstrated that LN18 cells expressed significant levels of FLAP and PPAR- alpha mRNAs. A low level of arachidonate 5-lipoxygenase (ALOX-5) mRNA was detected, but little, if any, arachidonate 12- lipoxygenase (ALOX-12) mRNA was present. In addition, MK886-induced apoptosis in LN18 cells was accompanied by a decrease in the protein and mRNA levels of vinculin, but not other focal adhesion proteins. In summary, the data presented here indicate that disruption of the actin-vinculin-cell-cytoskeleton matrix of the LN18 glioblastoma is a component of the MK886 induced apoptosis. In addition, MK886 treated LN18 cells could provide one model in which to investigate drugs that target lipoxygenase and PPAR-alpha pathways in the chemotherapeutic treatment of glioblastomas.

Efficacy of plant extracts against stored-products fungi

The fungistatic activity of six aqueous extracts of plants were tested against Aspergillus candidus, Aspergillus niger, Penicillium sp. and Fusarium culmorum. The plants were, chamomile (Anthemis nobilis L.), cinnamon (Cinnamomum verum J. Presl.), French lavender (Lavandula stoechas L.), garlic (Allium sativum L.), malva (Malva sylvestris L.) and peppermint (Mentha piperita L.). The more concentrated extracts of chamomile and malva inhibited totally the growth of the tested fungi with malva the most effective one(AU)

How reliable is axillary temperature measurement?

AIM: To assess whether axillary temperature measurements reliably reflect oral/rectal temperature measurements. METHODS: This observational study compared paired axillary-rectal and axillary-oral temperatures in a general paediatric ward with the participation of 225 children aged < or = 4 y and 112 children aged between 4 and 14 y. RESULTS: Changes in oral/rectal and axillary temperatures correlated significantly (p < 0.0001). However, axillary temperature measurements were significantly lower than both oral (mean -0.56 degrees C, SD 0.76 degrees C) and rectal measurements (0.38 degrees C; SD 0.76 degrees C). Ninety-five percent of axillary measurements fell within a 2.5-3 degrees C range around respective paired oral/rectal measurements. The mean difference increased with increasing temperature, and was 0.4 degrees C at low body temperatures, and over 1 degree C with a fever of 39 degrees C. Neither seasonal fluctuations nor the amount of clothing worn influenced this difference. CONCLUSION: Axillary temperatures in young children do not reliably reflect oral/rectal temperatures and should therefore be interpreted with caution.

Intraspecific variation in the venom electrophoretic profile of recently captured Crotalus durissus terrificus (Laurenti, 1768) snakes

The aim of this study was to compare individual venom samples of Crotalus durissus terrificus recently captured in the wild to evaluate possible differences in venom protein composition. Protein levels were quantified by biochemical method (Biuret) and then submitted to electrophoresis. Electrophoresis studies of native protein were performed in vertical slabs of polyacrylamide gel (PAGE), in an alkaline discontinuous buffer system, with a concentration of 10 per cent in the separation gel. SDS-PAGE was performed in PhastGel (8-25). Both gels were stained with Coomassie Blue. Gels were analyzed using the VDS-Pharmacia device. Our results indicate that all analyzed venom samples showed different protein composition, although common protein fractions were detected in some individual samples. Differences were observed between the different individual venom samples and so in the same specimen in relation to the time of collection, for both techniques used. Diet did not influence the variability of venom composition. There is a significant difference in native venom protein composition of males and females.

Evolutionary-derived anatomical characteristics and universal attractiveness.

Hominid fossils illustrate how modern humans have evolved anatomically. Included in the fossils are traits no longer phenotypically prevalent in humans (primitive) and phenotypic traits that have become increasingly prevalent (derived). In this study, published paleontological information about the anatomical evolution of humans was used to create line drawings of human form. Survey data were accumulated by having 759 individuals evaluate more than 40 anatomical traits. Each anatomical trait was presented as a panel of three line drawings intended to express the trait in a primitive, intermediate, and derived form. For each panel of three drawings, subjects were instructed to select the drawing they considered most attractive and then select the drawing they considered least attractive. The survey data indicate that males and females of diverse ages, races, cultures, and from varied geographical regions show commonality in their judgements of beauty of human form. The individuals surveyed appeared to have a strong aversion to primitive traits, preferring proportions and characteristics that are intermediate or more derived. In many instances, the evaluators preferred drawings that were exaggeratedly derived. The data may have relevance to the ongoing debate of whether averageness or atypicality is the essence of human beauty. Also, there was high agreement in judging the attractiveness of shapes and proportions in line drawings that were not immediately recognized to be representations of human form. These data could indicate that our general aesthetic sense for art, architecture, and fashion may be based on a subliminal reference to derived anatomical shapes and proportions. Overall, the data support the hypothesis that derived traits that are universally shared by anatomically modern humans may be the standard for our innate sense of beauty of human form.

Early treatment of the Class III malocclusion with rapid maxillary expansion and maxillary protraction.

This study comprised a sample of 31 patients with Class III malocclusion (21 girls and 10 boys), with ages ranging from 5 years 2 months to 11 years 6 months. All patients were in the deciduous or mixed dentition. The indicated treatment was rapid maxillary expansion, immediately followed by maxillary protraction with the facial mask. Mean treatment time was 8 months, varying from 4 to 24 months. The therapy induced both dental and skeletal alterations. Skeletal alterations consisted of maxillary anterior displacement and mandibular downward and backward rotation, improving facial profile. Dental alterations, known as dentoalveolar compensations, consisted of a tendency of labial tipping of the maxillary incisors and lingual tipping of the mandibular incisors.

Why Barbie is perceived as beautiful.

The long-term acceptance and success of the Barbie doll suggests the physical characteristics of the doll are perceived as attractive. When viewed in the context of universal attractiveness, response to the doll raises the question why Barbie is perceived as attractive. Published paleontological data on hominid fossils indicate how the shapes and anatomical proportions of humans have evolved. Included in the fossils are phenotypic traits no longer prevalent in humans (primitive) and phenotypic traits that have become increasingly prevalent (derived). It is noted that the anatomical proportions of the Barbie doll are exaggerated and emphasize derived characteristics. It was proposed that in the perception of human form, derived traits are perceived as attractive while primitive traits are perceived as unattractive. Drawings and photographs were utilized to survey reactions to a comparison of primitive vs derived traits by 495 subjects, instructed to select the shape or proportion they considered more attractive. There was significant agreement among the subjects that derived anatomical traits were perceived as more attractive than primitive ones. The Barbie doll is illustrative of how human beauty has evolved and indicates elements of human form that appear beautiful. The doll emphasizes our derived evolutionary traits and, possibly, that is why the doll is perceived as attractive.

Clinical features and hormonal characteristics in a case of ovarian arrhenoblastoma.

We report a case of a 34-year-old woman affected with ovarian arrhenoblastoma characterized by very high testosterone (T) levels (34.0-60.0 ng/ml; n.v.0.2-0.9) and suppressed gonadotropin levels. The physical examination revealed: severe hirsutism, acne, amenorrhea and other virilization signs. Basal hormonal evaluation also showed a markedly elevated 17-hydroxyprogesterone (17-OHP) and a mild delta 4 Androstenedione (A) and dehydroepiandrosterone sulfate (DHEAs) increase. ACTH test induced only slight changes in androgen secretion. By contrast, dexamethasone test greatly decreased A and DHEAs whereas T levels were only partially suppressed. Moreover, hCG test was clearly stimulatory for T and A. Suppressed gonadotropin levels did not respond to LHRH stimulation. The removal of the neoplasia was followed by normalization of T levels and increase of serum gonadotropins with subsequent restoration of a normal responsiveness to LHRH and resumption of an ovulatory menstrual cycle. This observation suggests that the high T levels played a primary role in the pathogenesis of the gonadotropin suppression and anovulation. Recovery of acne was complete whereas hirsutism score was reduced but still elevated after one year. This may be due to postoperative A and DHEAs levels slightly above the normal range, indicating the presence of adrenal hyperandrogenism.

Pubarche induction with testosterone treatment in women with panhypopituitarism.

OBJECTIVE: To evaluate the efficacy and safety of T therapy in the induction of pubic hair growth in women with congenital panhypopituitarism. DESIGN: Prospective clinical study. SETTING: Patients followed at the University Endocrinology Clinic. PATIENTS: Four women with congenital panhypopituitarism, showing no pubic hair development, currently treated with substitutive therapy with L-thyroxine, cortisone acetate, and estrogen-progestin combination. INTERVENTIONS: A long-acting T preparation (25 to 50 mg) was given IM each month; serum T levels were determined before and after 12 and 24 months of therapy. MAIN OUTCOME MEASURE: Evaluation of pubarche stages (according to Tanner classification of stages). RESULTS: Patients developed pubarche (Tanner stage 3 to 5) after 3 to 18 months of T therapy. Testosterone levels were within the normal range during treatment. No hirsutism or other side effects were recorded. CONCLUSION: A cautious T treatment represents an effective and safe approach to the problem of pubarche induction in women with congenital panhypopituitarism.

Effect of zinc supplementation on metallothionein, copper, and zinc concentration in various tissues of copper-loaded rats.

The present study was designed to investigate the effects of Zn administration on metallothionein concentrations in the liver, kidney, and intestine of copper-loaded rats. Male CD rats were fed a diet containing 12 mg Cu and 67 mg Zn/kg body wt. They were divided into either acute or chronic experimental protocols. Rats undergoing acute experiments received daily ip injections of either Cu (3 mg/kg body wt) or Zn (10 mg/kg body wt) for 3 d. Chronic experiments were carried out on rats receiving Cu ip injections on d 1, 2, 3, 10, 17, and 24, Cu injections plus a Zn-supplemented diet containing 5 g Zn/kg solid diet, or a Zn-supplemented diet alone. Rats injected Zn or Cu had increased MT concentrations in liver and kidney. Zn produced the most important effects and the liver was the most responsive organ. Rats fed a Zn-supplemented diet had significantly higher MT concentrations in liver and intestine with respect to controls. Increased MT synthesis in the liver may contribute to copper detoxification; the hypothesis of copper entrapment in enterocytes cannot be confirmed.

Collagen-induced rat platelet reactivity is enhanced in whole blood in both the presence and absence of dense granule secretion.

Collagen induced aggregation, ATP secretion and thromboxane (TxB2) generation of storage pool deficient platelets were compared to normal platelets of closely related rat strains. Platelet function was monitored in citrated-platelet-rich-plasma (PRP) and citrated whole blood. Wistar (W) and fawn-hooded (FH) rat strains and their F2 hybrids were utilized. The W strain, which is ancestral to the FH strain, is not storage pool deficient while the FH strain is. This was manifested by the total lack of collagen induced ATP secretion from platelets of the FH strain while the platelets of the W strain secreted normally. Utilizing platelets from the F2 generation of WxFH matings, the absence of dense granule secretion (ATP) from the FH platelets, as well as other platelet defects of FH rats, were shown to be associated with homozygosity for the red-eyed dilution gene [r]. The non-secreting FH platelets were utilized to determine the effects of secreted dense granule constituents upon collagen induced aggregation and TxB2 generation. The non-secreting storage pool deficient platelets did aggregate and did generate TxB2 upon stimulation with collagen; however, the storage pool deficient FH platelets demonstrated less TxB2 generation and did not aggregate as effectively as the normally secreting platelets of the W strain. When evaluating collagen induced platelet function in whole blood as compared to PRP, the storage pool deficient platelets remained less reactive than normally secreting platelets, but both platelet types demonstrated enhanced aggregation and increased TxB2 generation in whole blood.