Identification of candidate genes affecting Delta9-tetrahydrocannabinol biosynthesis in Cannabis sativa.

RNA isolated from the glands of a Delta(9)-tetrahydrocannabinolic acid (THCA)-producing strain of Cannabis sativa was used to generate a cDNA library containing over 100 000 expressed sequence tags (ESTs). Sequencing of over 2000 clones from the library resulted in the identification of over 1000 unigenes. Candidate genes for almost every step in the biochemical pathways leading from primary metabolites to THCA were identified. Quantitative PCR analysis suggested that many of the pathway genes are preferentially expressed in the glands. Hexanoyl-CoA, one of the metabolites required for THCA synthesis, could be made via either de novo fatty acids synthesis or via the breakdown of existing lipids. qPCR analysis supported the de novo pathway. Many of the ESTs encode transcription factors and two putative MYB genes were identified that were preferentially expressed in glands. Given the similarity of the Cannabis MYB genes to those in other species with known functions, these Cannabis MYBs may play roles in regulating gland development and THCA synthesis. Three candidates for the polyketide synthase (PKS) gene responsible for the first committed step in the pathway to THCA were characterized in more detail. One of these was identical to a previously reported chalcone synthase (CHS) and was found to have CHS activity. All three could use malonyl-CoA and hexanoyl-CoA as substrates, including the CHS, but reaction conditions were not identified that allowed for the production of olivetolic acid (the proposed product of the PKS activity needed for THCA synthesis). One of the PKS candidates was highly and specifically expressed in glands (relative to whole leaves) and, on the basis of these expression data, it is proposed to be the most likely PKS responsible for olivetolic acid synthesis in Cannabis glands

Location and course of the mandibular canal in dentate patients: morphometric study using cone-beam computed tomography

Background: The morphometric characteristics of the mandibular canal (MC) may vary according to the characteristics of the population studied. Correct location of the MC is fundamental for the indication and planning of different dental treatments, and it is therefore essential to have clinical parameters indicating its approximate location. The aim of this study was to describe the location and course of the MC by morphometric relations in the mandibular body, from the mental foramen to distal of the first molar, in dentate adult patients. Materials and methods: We analysed 55 cone-beam computed tomography (CBCT) of male and female patients, aged over 18 years and with fully dentate to the first molar. In each CBCT we selected five coronal sections (A–E) of the mandibular body at different levels using the teeth as references. We determined different morphometric measurements in each section to relate the MC with the corticals of the mandibular body (m1, m2, m3, m4), their orientation to lingual (F) and the thickness of the mandibular corticals (B1, B2 and B3). Results: The distance between the MC and the alveolar crests was greater in males than in females in all the sections. In general, in all the hemiarches, the MC courses away from the vestibular cortical of the mandible (and in some cases significantly towards the lingual cortical) from anterior to posterior in the mandibular body. In the left hemiarch of dentate females the MC describes a descent in the molar zone within the mandibular body. Any intervention in the mandible must be carried out with extreme caution to avoid damaging the vasculo-nervous bundle which passes through the MC. Conclusions: Cone-beam computed tomography is the best tool currently available for the planning and execution of surgical procedures, and is the only tool in clinical use which allows the precise course of the MC to be identified

Identification of candidate genes affecting Δ9-tetrahydrocannabinol biosynthesis in Cannabis sativa

RNA isolated from the glands of a Δ9-tetrahydrocannabinolic acid (THCA)-producing strain of Cannabis sativa was used to generate a cDNA library containing over 100 000 expressed sequence tags (ESTs). Sequencing of over 2000 clones from the library resulted in the identification of over 1000 unigenes. Candidate genes for almost every step in the biochemical pathways leading from primary metabolites to THCA were identified. Quantitative PCR analysis suggested that many of the pathway genes are preferentially expressed in the glands. Hexanoyl-CoA, one of the metabolites required for THCA synthesis, could be made via either de novo fatty acids synthesis or via the breakdown of existing lipids. qPCR analysis supported the de novo pathway. Many of the ESTs encode transcription factors and two putative MYB genes were identified that were preferentially expressed in glands. Given the similarity of the Cannabis MYB genes to those in other species with known functions, these Cannabis MYBs may play roles in regulating gland development and THCA synthesis. Three candidates for the polyketide synthase (PKS) gene responsible for the first committed step in the pathway to THCA were characterized in more detail. One of these was identical to a previously reported chalcone synthase (CHS) and was found to have CHS activity. All three could use malonyl-CoA and hexanoyl-CoA as substrates, including the CHS, but reaction conditions were not identified that allowed for the production of olivetolic acid (the proposed product of the PKS activity needed for THCA synthesis). One of the PKS candidates was highly and specifically expressed in glands (relative to whole leaves) and, on the basis of these expression data, it is proposed to be the most likely PKS responsible for olivetolic acid synthesis in Cannabis glands

Leprosy Associated with Atypical Cutaneous Leishmaniasis in Nicaragua and Honduras

In Central America, few cases of leprosy have been reported, but the disease may be unrecognized. Diagnosis is based on clinical criteria and histology. Preliminary field work in Nicaragua and Honduras found patients, including many children, with skin lesions clinically suggestive of atypical cutaneous leishmaniasis or indeterminate leprosy. Histology could not distinguish these diseases although acid-fast organisms were visible in a few biopsies. Lesions healed after standard antimicrobial therapy for leprosy. In the present study, patients, family members, and other community members were skin-tested and provided nasal swabs and blood samples. Biopsies were taken from a subgroup of patients with clinical signs of infection. Two laboratories analyzed samples, using local in-house techniques. Mycobacterium leprae, Leishmania spp. and Leishmania infantum were detected using polymerase chain reactions. Mycobacterium leprae DNA was detected in blood samples and nasal swabs, including some cases where leprosy was not clinically suspected. Leishmania spp. were also detected in blood and nasal swabs. Most biopsies contained Leishmania DNA and coinfection of Leishmania spp. with M. leprae occurred in 33% of cases. Mycobacterium leprae DNA was also detected and sequenced from Nicaraguan and Honduran environmental samples. In conclusion, leprosy and leishmaniasis are present in both regions, and leprosy appears to be widespread. The nature of any relationship between these two pathogens and the epidemiology of these infections need to be elucidated

Splicing Machinery is Dysregulated in Pituitary Neuroendocrine Tumors and is Associated with Aggressiveness Features

Pituitary neuroendocrine tumors (PitNETs) constitute approximately 15% of all brain tumors, and most have a sporadic origin. Recent studies suggest that altered alternative splicing and, consequently, appearance of aberrant splicing variants, is a common feature of most tumor pathologies. Moreover, spliceosome is considered an attractive therapeutic target in tumor pathologies, and the inhibition of SF3B1 (e.g., using pladienolide-B) has been shown to exert antitumor effects. Therefore, we aimed to analyze the expression levels of selected splicing-machinery components in 261 PitNETs (somatotropinomas/non-functioning PitNETS/corticotropinomas/prolactinomas) and evaluated the direct effects of pladienolide-B in cell proliferation/viability/hormone secretion in human PitNETs cell cultures and pituitary cell lines (AtT-20/GH3). Results revealed a severe dysregulation of splicing-machinery components in all the PitNET subtypes compared to normal pituitaries and a unique fingerprint of splicing-machinery components that accurately discriminate between normal and tumor tissue in each PitNET subtype. Moreover, expression of specific components was associated with key clinical parameters. Interestingly, certain components were commonly dysregulated throughout all PitNET subtypes. Finally, pladienolide-B reduced cell proliferation/viability/hormone secretion in PitNET cell cultures and cell lines. Altogether, our data demonstrate a drastic dysregulation of the splicing-machinery in PitNETs that might be associated to their tumorigenesis, paving the way to explore the use of specific splicing-machinery components as novel diagnostic/prognostic and therapeutic targets in PitNETs

A Self-Organizing Algorithm for Modeling Protein Loops

Protein loops, the flexible short segments connecting two stable secondary structural units in proteins, play a critical role in protein structure and function. Constructing chemically sensible conformations of protein loops that seamlessly bridge the gap between the anchor points without introducing any steric collisions remains an open challenge. A variety of algorithms have been developed to tackle the loop closure problem, ranging from inverse kinematics to knowledge-based approaches that utilize pre-existing fragments extracted from known protein structures. However, many of these approaches focus on the generation of conformations that mainly satisfy the fixed end point condition, leaving the steric constraints to be resolved in subsequent post-processing steps. In the present work, we describe a simple solution that simultaneously satisfies not only the end point and steric conditions, but also chirality and planarity constraints. Starting from random initial atomic coordinates, each individual conformation is generated independently by using a simple alternating scheme of pairwise distance adjustments of randomly chosen atoms, followed by fast geometric matching of the conformationally rigid components of the constituent amino acids. The method is conceptually simple, numerically stable and computationally efficient. Very importantly, additional constraints, such as those derived from NMR experiments, hydrogen bonds or salt bridges, can be incorporated into the algorithm in a straightforward and inexpensive way, making the method ideal for solving more complex multi-loop problems. The remarkable performance and robustness of the algorithm are demonstrated on a set of protein loops of length 4, 8, and 12 that have been used in previous studies

Exploring Health Science Students’ Notions on Organ Donation and Transplantation: A Multicenter Study

The knowledge acquired during university education about organ donation and transplantation (ODT) decisively influences the information future health professionals transmit. This is important in ODT where the participation of the general public is essential to obtain organs. Objective: To determine notions of Spanish medicine and nursing students on ODT and its relationship with attitude toward ODT. Methods and Design: and design. We conducted a sociologic, multicenter, and observational study. The population for our study consisted of medical and nursing students in Spanish universities. Our database was the Collaborative International Donor Project, stratified by geographic area and academic course. A validated questionnaire (PCID-DTO-RIOS) was self-administered and completed anonymously. Our sample consisted of 9598 medical and 10, 566 nursing students (99% confidence interval; precision of ±1%), stratified by geographic area and year of study. Results: The completion rate for our study was 90%. Only 20% (n=3640) of students thought their notions on ODT were good; 41% (n=7531) thought their notions were normal; 36% (n=6550) thought their notions were scarce. Comparing groups, there were differences between those who believed that their notions on ODT were good (44% nursing vs 56% medical students; P < .000), and those who believed it scarce (54% nursing vs 46% medical students; P < .000). Notions on ODT were related with attitude toward the donation of one''s own organs: those who considered their notions were good were more in favor then those who considered it scarce (88% vs 72%; P < .000). Conclusion: Only 20% of Spanish medical and nursing students thought their notions on ODT were good. Having good knowledge is related to a favorable attitude towards ODT. Receiving specific information on the subject could improve their knowledge about ODT during their training

Characteristics and Outcomes in Patients With COVID-19 and Acute Ischemic Stroke: The Global COVID-19 Stroke Registry

Recent case-series of small size implied a pathophysiological association between coronavirus disease 2019 (COVID-19) and severe large-vessel acute ischemic stroke. Given that severe strokes are typically associated with poor prognosis and can be very efficiently treated with recanalization techniques, confirmation of this putative association is urgently warranted in a large representative patient cohort to alert stroke clinicians, and inform pre- and in-hospital acute stroke patient pathways. We pooled all consecutive patients hospitalized with laboratory-confirmed COVID-19 and acute ischemic stroke in 28 sites from 16 countries. To assess whether stroke severity and outcomes (assessed at discharge or at the latest assessment for those patients still hospitalized) in patients with acute ischemic stroke are different between patients with COVID-19 and non-COVID-19, we performed 1:1 propensity score matching analyses of our COVID-19 patients with non-COVID-19 patients registered in the Acute Stroke Registry and Analysis of Lausanne Registry between 2003 and 2019. Between January 27, 2020, and May 19, 2020, 174 patients (median age 71.2 years; 37.9% females) with COVID-19 and acute ischemic stroke were hospitalized (median of 12 patients per site). The median National Institutes of Health Stroke Scale was 10 (interquartile range [IQR], 4-18). In the 1:1 matched sample of 336 patients with COVID-19 and non-COVID-19, the median National Institutes of Health Stroke Scale was higher in patients with COVID-19 (10 [IQR, 4-18] versus 6 [IQR, 3-14]), P=0.03; (odds ratio, 1.69 [95% CI, 1.08-2.65] for higher National Institutes of Health Stroke Scale score). There were 48 (27.6%) deaths, of which 22 were attributed to COVID-19 and 26 to stroke. Among 96 survivors with available information about disability status, 49 (51%) had severe disability at discharge. In the propensity score-matched population (n=330), patients with COVID-19 had higher risk for severe disability (median mRS 4 [IQR, 2-6] versus 2 [IQR, 1-4], P<0.001) and death (odds ratio, 4.3 [95% CI, 2.22-8.30]) compared with patients without COVID-19. Our findings suggest that COVID-19 associated ischemic strokes are more severe with worse functional outcome and higher mortality than non-COVID-19 ischemic strokes

Characteristics and Outcomes in Patients With COVID-19 and Acute Ischemic Stroke: The Global COVID-19 Stroke Registry.

Recent case-series of small size implied a pathophysiological association between coronavirus disease 2019 (COVID-19) and severe large-vessel acute ischemic stroke. Given that severe strokes are typically associated with poor prognosis and can be very efficiently treated with recanalization techniques, confirmation of this putative association is urgently warranted in a large representative patient cohort to alert stroke clinicians, and inform pre- and in-hospital acute stroke patient pathways. We pooled all consecutive patients hospitalized with laboratory-confirmed COVID-19 and acute ischemic stroke in 28 sites from 16 countries. To assess whether stroke severity and outcomes (assessed at discharge or at the latest assessment for those patients still hospitalized) in patients with acute ischemic stroke are different between patients with COVID-19 and non-COVID-19, we performed 1:1 propensity score matching analyses of our COVID-19 patients with non-COVID-19 patients registered in the Acute Stroke Registry and Analysis of Lausanne Registry between 2003 and 2019. Between January 27, 2020, and May 19, 2020, 174 patients (median age 71.2 years; 37.9% females) with COVID-19 and acute ischemic stroke were hospitalized (median of 12 patients per site). The median National Institutes of Health Stroke Scale was 10 (interquartile range [IQR], 4-18). In the 1:1 matched sample of 336 patients with COVID-19 and non-COVID-19, the median National Institutes of Health Stroke Scale was higher in patients with COVID-19 (10 [IQR, 4-18] versus 6 [IQR, 3-14]), P=0.03; (odds ratio, 1.69 [95% CI, 1.08-2.65] for higher National Institutes of Health Stroke Scale score). There were 48 (27.6%) deaths, of which 22 were attributed to COVID-19 and 26 to stroke. Among 96 survivors with available information about disability status, 49 (51%) had severe disability at discharge. In the propensity score-matched population (n=330), patients with COVID-19 had higher risk for severe disability (median mRS 4 [IQR, 2-6] versus 2 [IQR, 1-4], P&lt;0.001) and death (odds ratio, 4.3 [95% CI, 2.22-8.30]) compared with patients without COVID-19. Our findings suggest that COVID-19 associated ischemic strokes are more severe with worse functional outcome and higher mortality than non-COVID-19 ischemic strokes

Working relationships between obstetric care staff and their managers: a critical incident analysis

Background Malawi continues to experience critical shortages of key health technical cadres that can adequately respond to Malawi’s disease burden. Difficult working conditions contribute to low morale and frustration among health care workers. We aimed to understand how obstetric care staff perceive their working relationships with managers. Methods A qualitative exploratory study was conducted in health facilities in Malawi between October and December 2008. Critical Incident Analysis interviews were done in government district hospitals, faith-based health facilities, and a sample of health centres’ providing emergency obstetric care. A total of 84 service providers were interviewed. Data were analyzed using NVivo 8 software. Results Poor leadership styles affected working relationships between obstetric care staff and their managers. Main concerns were managers’ lack of support for staff welfare and staff performance, lack of mentorship for new staff and junior colleagues, as well as inadequate supportive supervision. All this led to frustrations, diminished motivation, lack of interest in their job and withdrawal from work, including staff seriously considering leaving their post. Conclusions Positive working relationships between obstetric care staff and their managers are essential for promoting staff motivation and positive work performance. However, this study revealed that staff were demotivated and undermined by transactional leadership styles and behavior, evidenced by management by exception and lack of feedback or recognition. A shift to transformational leadership in nurse-manager relationships is essential to establish good working relationships with staff. Improved providers’ job satisfaction and staff retentionare crucial to the provision of high quality care and will also ensure efficiency in health care delivery in Malawi