Three Undescribed Iridoid Derivatives Isolated from the Fruits of Vitex trifolia with Their Cytotoxic and Antimicrobial Activities.

Vitex trifolia L. is a medicinal plant and widely distributed in the northern mountainous areas of Vietnam. Phytochemical study on the fruits of this plant led to the isolation of nine iridoid derivatives (1-9) including three undescribed compounds (1-3). Their structures were elucidated to be 3''-hydroxyscrophuloside A1 (1), 3''-hydroxycallicoside D (2), 2'-p-hydroxybenzoylaucubin (3), 6'-p-hydroxybenzoylmussaenosidic acid (4), nishindaside (5), agnuside (6), 10-O-vanilloylaucubin (7), 6'-O-p-hydroxybenzoyl-gardoside (8), and buddlejoside B (9) based on extensive analyses of HR-ESI-MS, 1D and 2D NMR spectra. Compounds 1, 2, 4, and 8 significantly posessed anti-barterial activity against Enterococcus faecalis, Staphylococcus aureus, Escherichia coli, Pseudomonas aeruginosa strains with MIC values in range of 16-64 µg/mL. At concentration of 20 µM, compounds 1-9 did not show cytotoxic effects against human lung cancer cells (PC9).

Diffuse Intracerebral Hemorrhage in an Infant With a Novel Homozygous Variant Leading to Severe Protein C Deficiency.

Protein C is a circulating anticoagulant that inhibits factor Va and VIIIa and promotes fibrinolysis. Compound heterozygous or homozygous variants in the Protein C gene (PROC) lead to severe deficiency of protein C and affected neonates typically present shortly after birth with purpura fulminans. We describe an infant who suffered a diffuse intracranial hemorrhage as a neonate and presented with purpura fulminans as an older infant which led to investigations that were consistent with severe protein C deficiency. We demonstrate subacute findings on neuroimaging and suggest this condition should be considered with neonatal presentations of bilateral intraparenchymal hemorrhage.

Hb Calgary (HBB: c.194G>T): A Highly Unstable Hemoglobin Variant with a ß-Thalassemia Major Phenotype.

We describe two unrelated patients, both heterozygous for an unstable hemoglobin (Hb) variant named Hb Calgary (HBB: c.194G>T) that causes severe hemolytic anemia and dyserythorpoietic, resulting in transfusion dependence and iron overload. The molecular pathogenesis is a missense variation on the ß-globin gene, presumed to lead to an unstable Hb. The phenotype of Hb Calgary is particularly severe presenting as transfusion-dependent anemia in early infancy, precluding phenotypic diagnosis and highlighting the importance of early genetic testing in order to make an accurate diagnosis.

Fludarabine, Campath, and Low-Dose Cyclophosphamide (FCClow) with or without TBI Conditioning Results in Excellent Transplant Outcomes in Children with Severe Aplastic Anemia.

Various reduced-intensity conditioning regimens are in use for allogeneic hematopoietic cell transplant (HSCT) in patients with idiopathic severe aplastic anemia (SAA). We describe the use of fludarabine, Campath, and low-dose cyclophosphamide (FCClow) conditioning in 15 children undergoing related or unrelated donor transplants. Total body irradiation (TBI) of 2 Gy was added for unrelated donor HSCT. At a median follow-up of 2.3 years, the failure-free survival was 100%, with low rates of infection and toxicity. There was no occurrence of grade III to IV acute graft-versus-host disease (GVHD). All patients had full donor myeloid chimerism post-HSCT, even with mixed chimerism in the T cell lineage. The absence of chronic GVHD and long-term stable mixed donor T cell chimerism confirms immune tolerance following FCClow (± TBI) conditioned transplantation in children with SAA.

Interactions between Curcumin Derivatives and Amyloid-ß Fibrils: Insights from Molecular Dynamics Simulations.

The aggregation of amyloid-ß (Aß) peptides into senile plaques is a hallmark of Alzheimer's disease (AD) and is hypothesized to be the primary cause of AD related neurodegeneration. Previous studies have shown the ability of curcumin to both inhibit the aggregation of Aß peptides into oligomers or fibrils and reduce amyloids in vivo. Despite the promise of curcumin and its derivatives to serve as diagnostic, preventative, and potentially therapeutic AD molecules, the mechanism by which curcumin and its derivatives bind to and inhibit Aß fibrils' formation remains elusive. Here, we investigated curcumin and a set of curcumin derivatives in complex with a hexamer peptide model of the Aß1-42 fibril using nearly exhaustive docking, followed by multi-ns molecular dynamics simulations, to provide atomistic-detail insights into the molecules' binding and inhibitory properties. In the vast majority of the simulations, curcumin and its derivatives remain firmly bound in complex with the fibril through primarily three different principle binding modes, in which the molecules interact with residue domain 17LVFFA21, in line with previous experiments. In a small subset of these simulations, the molecules partly dissociate the outermost peptide of the Aß1-42 fibril by disrupting ß-sheets within the residue domain 12VHHQKLVFF20. A comparison between binding modes leading or not leading to partial dissociation of the outermost peptide suggests that the latter is attributed to a few subtle key structural and energetic interaction-based differences. Interestingly, partial dissociation appears to be either an outcome of high affinity interactions or a cause leading to high affinity interactions between the molecules and the fibril, which could partly serve as a compensation for the energy loss in the fibril due to partial dissociation. In conjunction with this, we suggest a potential inhibition mechanism of Αß1-42 aggregation by the molecules, where the partially dissociated 16KLVFF20 domain of the outermost peptide could either remain unstructured or wrap around to form intramolecular interactions with the same peptide's 29GAIIG33 domain, while the molecules could additionally act as a patch against the external edge of the second outermost peptide's 16KLVFF20 domain. Thereby, individually or concurrently, these could prohibit fibril elongation.

Severe Congenital Protein C Deficiency: Practical Aspects of Management.

Subcutaneous (SC) protein C (PC) was used in a child with purpura fulminans secondary to severe congenital PC deficiency. For maintenance, PC 80-120 IU/kg, given over 60-90 min SC Q48hr, has been successful as a home therapy for more than 3 years. The treatment was monitored by measuring trough PC chromogenic activity (target ≥15%) and D-dimer levels. No change in clinical course was appreciated after discontinuing enoxaparin (and leaving the patient on prophylactic PC replacement alone). A significant discrepancy between clotting-based and chromogenic-based PC activity is shown.

Vitamin B12 Deficiency in Infancy: The Case for Screening.

The classic principles put forth by Wilson and Jungner are often applied to determine the suitability of a condition for universal newborn screening. The three cases described here portray the harmful effects of vitamin B12 deficiency in infancy. The challenges and opportunities of early recognition and treatment are highlighted. Screening newborns would allow early detection and prevention of severe neurological damage in vitamin B12 -deficient infants and enable diagnosis of unrecognized maternal pernicious anemia in asymptomatic mothers. However, lack of standardized methodology and screening cutoffs present challenges to the use of current tandem mass spectrometry technologies for screening.

Adhesion of the genome-sequenced Lactococcus lactis subsp. cremoris IBB477 strain is mediated by specific molecular determinants.

Understanding the nature of mucus-microbe interactions will provide important information that can help to elucidate the mechanisms underlying probiotic adhesion. This study focused on the adhesive properties of the Lactococcus lactis subsp. cremoris IBB477 strain, previously shown to persist in the gastrointestinal tract of germ-free rats. The shear flow-induced detachment of L. lactis cells was investigated under laminar flow conditions. Such a dynamic approach demonstrated increased adhesion to bare and mucin-coated polystyrene for IBB477, compared to that observed for the MG1820 control strain. To identify potential genetic determinants giving adhesive properties to IBB477, the improved high-quality draft genome sequence comprising chromosome and five plasmids was obtained and analysed. The number of putative adhesion proteins was determined on the basis of surface/extracellular localisation and/or the presence of adhesion domains. To identify proteins essential for the IBB477 specific adhesion property, nine deletion mutants in chromosomal genes have been constructed and analysed using adhesion tests on bare polystyrene as well as mucin-, fibronectin- or collagen IV-coated polystyrene plates in comparison to the wild-type strain. These experiments demonstrated that gene AJ89_07570 encoding a protein containing DUF285, MucBP and four Big_3 domains is involved in adhesion to bare and mucin-coated polystyrene. To summarise, in the present work, we characterised the adhesion of IBB477 under laminar flow conditions; identified the putative adherence factors present in IBB477, which is the first L. lactis strain exhibiting adhesive and mucoadhesive properties to be sequenced and demonstrated that one of the proteins containing adhesion domains contributes to adhesion.

Response and resistance to MEK inhibition in leukaemias initiated by hyperactive Ras.

The cascade comprising Raf, mitogen-activated protein kinase kinase (MEK) and extracellular signal-regulated kinase (ERK) is a therapeutic target in human cancers with deregulated Ras signalling, which includes tumours that have inactivated the Nf1 tumour suppressor. Nf1 encodes neurofibromin, a GTPase-activating protein that terminates Ras signalling by stimulating hydrolysis of Ras-GTP. We compared the effects of inhibitors of MEK in a myeloproliferative disorder (MPD) initiated by inactivating Nf1 in mouse bone marrow and in acute myeloid leukaemias (AMLs) in which cooperating mutations were induced by retroviral insertional mutagenesis. Here we show that MEK inhibitors are ineffective in MPD, but induce objective regression of many Nf1-deficient AMLs. Drug resistance developed because of outgrowth of AML clones that were present before treatment. We cloned clone-specific retroviral integrations to identify candidate resistance genes including Rasgrp1, Rasgrp4 and Mapk14, which encodes p38alpha. Functional analysis implicated increased RasGRP1 levels and reduced p38 kinase activity in resistance to MEK inhibitors. This approach represents a robust strategy for identifying genes and pathways that modulate how primary cancer cells respond to targeted therapeutics and for probing mechanisms of de novo and acquired resistance.

Three new species of the genus Loxoconcha (Crustacea, Ostracoda, Podocopida) from the Okinawa Islands, southern Japan.

Three new species of Ostracoda, Loxoconcha noharai sp. nov., L. santosi sp. nov. and L. sesokoensis sp. nov., are described from the Okinawa Islands, southern Japan. The two species Loxoconcha noharai sp. nov. and L. santosi sp. nov. live in estuaries, whereas the species L. sesokoensis sp. nov. lives in coral reefs. These species can be easily distinguished from other previously described Loxoconcha species by their morphological differences, mainly in the male copulatory organ, and distribution pattern of their pore systems. In addition, L. sesokoensis sp. nov. is suggested to be phylogenetically apart from any other Loxoconcha species which have been reported so far from Japan and the adjacent seas.

Subunit Interactions and cooperativity in the microtubule-severing AAA ATPase spastin.

Spastin is a hexameric ring AAA ATPase that severs microtubules. To see whether the ring complex funnels the energy of multiple ATP hydrolysis events to the site of mechanical action, we investigate here the cooperativity of spastin. Several lines of evidence indicate that interactions among two subunits dominate the cooperative behavior: (i) the ATPase activity shows a sigmoidal dependence on the ATP concentration; (ii) ATPγS displays a mixed-inhibition behavior for normal ATP turnover; and (iii) inactive mutant subunits inhibit the activity of spastin in a hyperbolic dependence, characteristic for two interacting species. A quantitative model based on neighbor interactions fits mutant titration experiments well, suggesting that each subunit is mainly influenced by one of its neighbors. These observations are relevant for patients suffering from SPG4-type hereditary spastic paraplegia and explain why single amino acid exchanges lead to a dominant negative phenotype. In severing assays, wild type spastin is even more sensitive toward the presence of inactive mutants than in enzymatic assays, suggesting a weak coupling of ATPase and severing activity.

Recent advances in computerized imaging and its vital roles in liver disease diagnosis, preoperative planning, and interventional liver surgery: A review.

The earliest and most accurate detection of the pathological manifestations of hepatic diseases ensures effective treatments and thus positive prognostic outcomes. In clinical settings, screening and determining the extent of a pathology are prominent factors in preparing remedial agents and administering appropriate therapeutic procedures. Moreover, in a patient undergoing liver resection, a realistic preoperative simulation of the subject-specific anatomy and physiology also plays a vital part in conducting initial assessments, making surgical decisions during the procedure, and anticipating postoperative results. Conventionally, various medical imaging modalities, e.g., computed tomography, magnetic resonance imaging, and positron emission tomography, have been employed to assist in these tasks. In fact, several standardized procedures, such as lesion detection and liver segmentation, are also incorporated into prominent commercial software packages. Thus far, most integrated software as a medical device typically involves tedious interactions from the physician, such as manual delineation and empirical adjustments, as per a given patient. With the rapid progress in digital health approaches, especially medical image analysis, a wide range of computer algorithms have been proposed to facilitate those procedures. They include pattern recognition of a liver, its periphery, and lesion, as well as pre- and postoperative simulations. Prior to clinical adoption, however, software must conform to regulatory requirements set by the governing agency, for instance, valid clinical association and analytical and clinical validation. Therefore, this paper provides a detailed account and discussion of the state-of-the-art methods for liver image analyses, visualization, and simulation in the literature. Emphasis is placed upon their concepts, algorithmic classifications, merits, limitations, clinical considerations, and future research trends.

Case report: Blotchy skin in a puffy neonate: is there a new association?

Introduction: Purpura fulminans in the neonatal population is a rare but potentially life-threatening condition complicated by thrombosis, resultant vital organ necrosis, and gangrene of the extremities. Considering the rapid evolution of the pathogenetic mechanism, an index of suspicion, early identification, and prompt intervention are imperative for improved outcomes. The majority of purpura fulminans cases have an infectious etiology, but it is essential to consider other congenital and acquired causes. Case description: We present a clinical case of a female neonate to emphasize the correlation between purpura fulminans, congenital chylothorax, involvement of the PAK2 gene, and the occurrence of retinal detachment in both eyes. After draining the congenital chylothorax, the neonate developed purpura fulminans due to a loss of protein C, S, and antithrombin factors, previously not reported in the literature. The purpuric lesions resolved after the administration of fresh frozen plasma. Subsequently, no recurring purpura fulminans lesions were noted following the normalization of the antithrombotic factor levels in the serum. Subsequently, the child also developed retinal detachment in both eyes.

Lenient rate control versus strict rate control for atrial fibrillation: a statistical analysis plan for the Danish Atrial Fibrillation (DanAF) randomized clinical trial.

BACKGROUND: A key decision in the treatment of atrial fibrillation is choosing between a rhythm control strategy or a rate control strategy as the main strategy. When choosing rate control, the optimal heart rate target is uncertain. The Danish Atrial Fibrillation trial is a randomized, multicenter, two-group, superiority trial comparing strict rate control versus lenient rate control in patients with either persistent or permanent atrial fibrillation at inclusion. To prevent bias arising from selective reporting and data-driven analyses, we developed a predefined description of the statistical analysis. METHODS: The primary outcome of this trial is the physical component score of the SF-36 questionnaire. A total of 350 participants will be enrolled based on a minimal important difference of 3 points on the physical component score of the SF-36 questionnaire, a standard deviation of 10 points, a statistical power of 80% (beta of 20%), and an acceptable risk of type I error of 5%. All secondary, exploratory, and echocardiographic outcomes will be hypothesis-generating. The analyses of all outcomes will be based on the intention-to-treat principle. We will analyze continuous outcomes using linear regression adjusting for "site," type of atrial fibrillation at inclusion (persistent/ permanent), left ventricular ejection fraction (≥ 40% or < 40%), and the baseline value of the outcome (all as fixed effects). We define our threshold for statistical significance as a p-value of 0.05 and assessments of clinical significance will be based on the anticipated intervention effects defined in the sample size and power estimations. Thresholds for both statistical and clinical significance will be assessed according to the 5-step procedure proposed by Jakobsen and colleagues. DISCUSSION: This statistical analysis plan will be published prior to enrolment completion and before any data are available and is sought to increase the validity of the DANish Atrial Fibrillation trial. TRIAL REGISTRATION: Clinicaltrials.gov NCT04542785. Registered on Sept 09, 2020.

Real-time investigation of the muco-adhesive properties of Lactococcus lactis using a quartz crystal microbalance with dissipation monitoring.

This work was devoted to probe, at the entire population level, interactions between mucins and Lactococcus lactis, using QCM-D. Real-time monitoring of adsorption on polystyrene of PGM (Pig Gastric Mucin) and subsequent adhesion of L. lactis was performed for IBB477 and MG1820 strains. Measuring simultaneously shifts in resonance frequency and dissipation on the polystyrene-coated crystal demonstrated a two-phase process for PGM adsorption. XPS analysis confirmed the presence of adsorbed mucin. The Voigt-based model was used to describe the QCM-D outputs. The predicted thickness of the PGM layer was consistent with the AFM experimental value. Adhesion of L. lactis to bare or PGM-coated polystyrene was then monitored, in combination with DAPI cell counting. Positive frequency shifts were caused by adhering bacteria. The presence of adsorbed PGM strongly reduced bacterial adhesion. However, adhesion of IBB477 to the PGM coating was greatly increased in comparison with that of MG1820. Muco-adhesion may be a highly variable and valuable phenotypic trait among L. lactis strains.

Functional outcomes and complications of tibial lengthening using unilateral external fixation and then plating. A prospective case series.

INTRODUCTION: and Importance: The purpose of this study was to assess the functional outcomes and complications of tibial lengthening using unilateral external fixation and then plating. MATERIAL AND METHODS: This was a prospective case series study that enrolled people of short stature or leg length discrepancy of more than 3 cm from January 2019 to January 2021. A total of 11 patients (one male and 10 females) were recruited for the study, including seven short statures and four patients with leg length discrepancies. The external fixaters in this study were Muller's frame or Nhan's frame. RESULTS: The average age of patients at the time of surgery was 25.89 years (range: 13-41 years). The study included 18 tibias that were lengthened and then plated. Average tibial lengthening was 6.89 ± 1.25 cm (21.87 ± 6.59%). The functional result was excellent in seven patients and good in four patients. Pin-track infection occurred in three tibias. There was one case of superficial infection. 12 legs (66.7%) developed ankle equinus after removing the external fixator. Four legs with severe equinus deformity were treated with percutaneous tendo-Achilles lengthening. Valgus deviation occurred in eight tibias. Peroneal nerve neuropraxia occurred in two legs during distraction. Distal migration of the fibula head occurred in four legs. CONCLUSION: Our study suggested that tibial lengthening using Nhan's external fixater or Muller frame then plating was safe and effective provided complications were looked for and kept in check. Equinus contracture, pin-site infection, and valgus alignment were the most common complications. LEVEL OF EVIDENCE: Level IV, prospective case series study.

Measuring kinetic dissociation/association constants between Lactococcus lactis bacteria and mucins using living cell probes.

In this work we focused on quantifying adhesion between Lactococcus lactis, the model for lactic acid bacteria (LAB) and mucins. Interactions between two strains of L. lactis (IBB477 and MG1820 as control) and pig gastric mucin-based coating were measured and compared with the use of atomic force microscopy. Analysis of retraction force-distance curves shed light on the differential contributions of nonspecific and specific forces. An increased proportion of specific adhesive events was obtained for IBB477 (20% vs. 5% for the control). Blocking assays with free pig gastric mucin and its O-glycan moiety showed that oligosaccharides play a major (but not exclusive) role in L. lactis-mucins interactions. Specific interactions were analyzed in terms of kinetic constants. An increase in the loading rate of atomic force microscope tip led to a higher force between interacting biological entities, which was directly linked to the kinetic dissociation constant (K(off)). Enhancing the contact time between the tip and the sample allowed an increase in the interaction probability, which can be related to the kinetic association constant (K(on)). Variations in the loading rate and contact time enabled us to determine K(on) (3.3 × 10(2) M(-1)·s(-1)) and K(off) (0.46 s(-1)), and the latter was consistent with values given in the literature for sugar-protein interactions.

Fibula lengthening then centralization for the treatment of pseudoarthrosis at the middle third of tibia with large leg-length discrepancy - A case report.

INTRODUCTION AND IMPORTANCE: Tibial dysplastic pseudoarthrosis associated with large leg-length discrepancy and extensive scarring was a great challenge for orthopedic surgeons. The treatment for these lesions was not defined. CASE PRESENTATION: We report a 24-year-old case of dysplastic pseudoarthrosis at the middle third of the right tibia with a 10 cm leg-length discrepancy and a 250 medial deviation and 200 internal torsion due to osteomyelitis 14 years previously. She was treated by fibula lengthening then centralization for the treatment of tibial pseudoarthrosis. Eight months after the operation, the patient reported no pain and became capable of walking without an orthosis. X-rays showed full bone union at the proximal tibiofibular synostosis and the lengthening site of the right fibula. The distal tibiofibular synostosis was healing. The range of movement of the knee and ankle was restored. CLINICAL DISCUSSION: This technique was different from the Huntington procedure. A good union between the transposed fibula and the remaining part of the fibula brought a better strength of the reconstructed tibia. The tibial pseudoarthrosis, leg-length discrepancy, and malalignment had been solved by our techniques. The procedure was easy, rapid, and inexpensive without requiring microsurgery skills and special devices. CONCLUSION: Leg lengthening then centralization of ipsilateral fibular graft is an excellent option for reconstruction of a tibial pseudarthrosis with a large leg-length discrepancy. It is a safe, rapid, and inexpensive procedure. The procedure was not difficult, and microvascular skills and special devices were not required. LEVEL OF EVIDENCE: A case report.