Explore the vast range of titles available.

The morphogen Decapentaplegic (Dpp) has been implicated in both wing patterning and growth in fruitflies; here, a CRISPR–Cas9 approach is used to rule out the Dpp gradient driving wing growth. Morphogen patterning in Drosophila In Drosophila , the morphogen Decapentaplegic (Dpp), which is homologous to bone morphogenetic protein, has been implicated both in wing patterning and growth. Dpp is secreted from a central stripe in developing wing, and forms a gradient that is thought to be essential for its role in patterning. How the Dpp gradient drives proliferation across the whole tissue has been a matter of debate, joined by two groups reporting in this issue of Nature . Stefan Harmansa et al . have developed a morphotrap approach that relies on the expression of a membrane-tethered GFP antibody to immobilize GFP-tagged Dpp in a specific region of the wing. They show that although the absence of spreading disrupts the patterning of the wing, lateral cells still divide normally, ruling out a role for the Dpp gradient in regulating lateral wing growth. Takuya Akiyama and Matthew Gibson have used a CRISPR–Cas9-mediated approach to ablate Dpp expression specifically in the stripe and although the resulting animals exhibit patterning defects, their cell proliferation and growth remained relatively normal. This rules out a role for the Dpp stripe in modulating wing growth. As a central model for morphogen action during animal development, the bone morphogenetic protein 2/4 (BMP2/4)-like ligand Decapentaplegic (Dpp) is proposed to form a long-range signalling gradient that directs both growth and pattern formation during Drosophila wing disc development 1 , 2 , 3 , 4 , 5 , 6 . While the patterning role of Dpp secreted from a stripe of cells along the anterior–posterior compartmental boundary is well established 1 , 2 , 6 , the mechanism by which a Dpp gradient directs uniform cell proliferation remains controversial and poorly understood 7 , 8 , 9 , 10 , 11 , 12 , 13 . Here, to determine the precise spatiotemporal requirements for Dpp during wing disc development, we use CRISPR–Cas9-mediated genome editing to generate a flippase recognition target ( FRT )-dependent conditional null allele. By genetically removing Dpp from its endogenous stripe domain, we confirm the requirement of Dpp for the activation of a downstream phospho-Mothers against dpp (p-Mad) gradient and the regulation of the patterning targets spalt ( sal ), optomotor blind ( omb ; also known as bifid ) and brinker ( brk ). Surprisingly, however, third-instar wing blade primordia devoid of compartmental dpp expression maintain relatively normal rates of cell proliferation and exhibit only mild defects in growth. These results indicate that during the latter half of larval development, the Dpp morphogen gradient emanating from the anterior–posterior compartment boundary is not directly required for wing disc growth.

The basement membrane (BM) plays critical roles in stem cell maintenance and activity control. Here we show that chondroitin sulfate (CS), a major component of the Drosophila midgut BM, is required for proper control of intestinal stem cells (ISCs). Loss of Chsy , a critical CS biosynthetic gene, resulted in elevated levels of ISC proliferation during homeostasis, leading to midgut hyperplasia. Regeneration assays demonstrated that Chsy mutant ISCs failed to properly downregulate mitotic activity at the end of regeneration. We also found that CS is essential for the barrier integrity to prevent leakage of the midgut epithelium. CS is known to be polymerized by the action of the complex of Chsy and another critical protein, Chondroitin polymerizing factor (Chpf). We found that Chpf mutants show increased ISC division during midgut homeostasis and regeneration, similar to Chsy mutants. As Chpf is induced by a tissue damage during regeneration, our data suggest that Chpf functions with Chsy to facilitate CS remodeling and stimulate tissue repair. We propose that the completion of the repair of CS-containing BM acts as a prerequisite to properly terminate the regeneration process.

Stool consistency is evaluated mainly in reference to indirect indicators such as water content or the appearance of stool forms using Bristol Stool Form Scale (BSFS). Methods of measurement are limited. We thus aimed to develop a simple protocol for direct measurement of stool consistency using the TA.XTExpress Texture Analyser (Stable Micro Systems Ltd.). We developed a protocol which enables mechanical quantification of the gram-force against a cylindrical probe (ø 6 mm) pushed into the stool surface at 2.0 mm/s to 5 mm depth. The consistency of 252 stools collected from 40 healthy Belgians was evaluated by the direct method and by the indirect indicators (water content and BSFS) for comparison. The log-transformed stool consistency values measured by the texture analyzer had a negative linear correlation with the stool water contents ( r rm  = − 0.781) with homoscedastic variance, suggesting the appropriateness of the new protocol. They showed a similar correlation with the BSFS, but with a large variance in the consistency values of normal stool forms. This correlation was much smaller for BSFS scored by subjects ( r rm  = − 0.587) than by experts ( r rm  = − 0.789), collectively indicating BSFS as a rough indicator of stool consistency susceptible to subjective bias despite its effectiveness in clinical use. The optimized direct method using the texture analyzer enables the accurate quantification of stool consistency, which facilitates understanding of the intestinal environment and function and thus may enhance the value of the stool as a predictor of human health.

Purpose To evaluate the effectiveness of MicroPulse ® transscleral laser therapy (MP-TLT) using both the original and revised MicroPulse P3 ® delivery probes in patients with refractory glaucoma. Study design Retrospective study Methods We analyzed the data of patients with refractory glaucoma who underwent MP-TLT with the original and with the revised MicroPulse P3 ® Delivery Device from January 2018 to December 2020 at the University of Tokyo Hospital and who were followed up for 3 months. Patients’ demographics, preoperative and postoperative intraocular pressure (IOP), medication scores, best-corrected visual acuity (BCVA), and complications were analyzed. Results This study enrolled 40 patients in both probe groups. At baseline, the mean IOP was 31.1 mmHg in the original group and 29.2 mmHg in the revised group, and it decreased to 22.6 mmHg and 21.6 mmHg after 3 months, respectively (paired t-test p=0.001, 0.001). No significant difference was noted between the number of medications and BCVA at baseline and 3 months in either probe group (paired t-test, p>0.05). Nor were any serious complications observed. Conclusion MP-TLT using the revised MicroPulse P3 ® Delivery Device for the treatment of refractory glaucoma was effective in reducing IOP across glaucoma types with an excellent safety profile and was well-tolerated by the patients.

Although finely divided ball-milled whole cell walls do not completely dissolve in dimethylsulfoxide (DMSO), they readily swell producing a gel. Solution-state two-dimensional (2D) nuclear magnetic resonance (NMR) of this gel, produced directly in the NMR tube, provides an interpretable structural fingerprint of the polysaccharide and lignin components of the wall without actual solubilization, and without structural modification beyond that inflicted by the ball milling and ultrasonication steps. Since the cellulose is highly crystalline and difficult to swell, the component may be under-represented in the spectra. The method however provides a more rapid method for comparative structural evaluation of plant cell walls than is currently available. With the new potential for chemometric analysis using the 2D NMR fingerprint, this method may find application as a secondary screen for selecting biomass lines and for optimizing biomass processing and conversion efficiencies.

Endoplasmic reticulum (ER) stress is caused by accumulation of unfolded and misfolded proteins in the ER, thereby compromising its vital cellular functions in protein production and secretion. Genome wide association studies in humans as well as experimental animal models linked ER stress in intestinal epithelial cells (IECs) with intestinal disorders including inflammatory bowel diseases. However, the mechanisms linking the outcomes of ER stress in IECs to intestinal disease have not been clarified. In this study, we investigated the impact of ER stress on intestinal epithelial barrier function using human colon carcinoma-derived Caco-2 monolayers. Tunicamycin-induced ER stress decreased the trans-epithelial electrical resistance of Caco-2 monolayers, concomitant with loss of cellular plasma membrane integrity. Epithelial barrier disruption in Caco-2 cells after ER stress was not caused by caspase- or RIPK1-dependent cell death but was accompanied by lysosomal rupture and up-regulation of the ER stress markers Grp78, sXBP1 and Chop. Interestingly, several bifidobacteria species inhibited tunicamycin-induced ER stress and thereby diminished barrier disruption in Caco-2 monolayers. Together, these results showed that ER stress compromises the epithelial barrier function of Caco-2 monolayers and demonstrate beneficial impacts of bifidobacteria on ER stress in IECs. Our results identify epithelial barrier loss as a potential link between ER stress and intestinal disease development, and suggest that bifidobacteria could exert beneficial effects on this phenomenon.

Background The importance of the gut microbiota at the early stage of life and their longitudinal effect on host health have recently been well investigated. In particular, Bifidobacterium longum subsp. longum , a common component of infant gut microbiota, appears in the gut shortly after birth and can be detected there throughout an individual’s lifespan. However, it remains unclear whether this species colonizes in the gut over the long term from early infancy. Here, we investigated the long-term colonization of B . longum subsp. longum by comparing the genotypes of isolates obtained at different time points from individual subjects. Strains were isolated over time from the feces of 12 subjects followed from early infancy (the first six months of life) up to childhood (approximately six years of age). We also considered whether the strains were transmitted from their mothers’ perinatal samples (prenatal feces and postnatal breast milk). Results Intra-species diversity of B. longum subsp. longum was observed in some subjects’ fecal samples collected in early infancy and childhood, as well as in the prenatal fecal samples of their mothers. Among the highlighted strains, several were confirmed to colonize and persist in single individuals from as early as 90 days of age for more than six years; these were classified as long-term colonizers. One of the long-term colonizers was also detected from the corresponding mother’s postnatal breast milk. Quantitative polymerase chain reaction data suggested that these long-term colonizers persisted in the subjects’ gut despite the existence of the other predominant species of Bifidobacterium . Conclusions Our results showed that several strains belonging to B. longum subsp. longum colonized in the human gut from early infancy through more than six years, confirming the existence of long-term colonizers from this period. Moreover, the results suggested that these strains persisted in the subjects’ gut while co-existing with the other predominant bifidobacterial species. Our findings also suggested the importance of microbial-strain colonization in early infancy relative to their succession and showed the possibility that probiotics targeting infants might have longitudinal effects. Trial Registration TRN: ISRCTN25216339 . Date of registration: 11/03/2016. Prospectively registered.

Background The neutrophil-to-lymphocyte ratio (NLR), an inflammatory marker, is known to predict postoperative complications. However, its predictive value for postoperative inflammatory status and postoperative pain (POP) following cardiac surgery remains unclear. This study aimed to determine whether elevated preoperative NLR is associated with increased postoperative analgesic consumption. Methods We retrospectively analyzed patients aged over 18 years who underwent cardiac surgery and received intravenous patient-controlled analgesia (IVPCA) for POP between April 2017 and March 2019. The IVPCA solution contained fentanyl (5 μg/mL), lidocaine (8 mg/mL), ketamine (0.8 mg/mL), and droperidol (50 μg/mL). The primary outcome was the correlation between preoperative NLR and postoperative IVPCA consumption. Patients were categorized into high (> median) and low (< median) preoperative NLR groups. To adjust for baseline differences, propensity score matching was applied. Post-matching, we compared perioperative variables and reassessed the association between preoperative NLR and IVPCA usage. Results A total of 131 patients were included. The median preoperative NLR was 2.29 (IQR: 1.58-3.39). Across all patients, preoperative NLR was not correlated with postoperative IVPCA consumption (ρ = -0.08, P = 0.34). After excluding the patient with the median NLR, 76 patients (38 in each group) remained for analysis post-matching. IVPCA consumption was comparable between the high and low NLR groups (108.6 (77.6-164.0) mL vs. 97.1 (83.1-148.0) mL, P = 0.97). However, patients with higher preoperative NLR had significantly elevated NLR on postoperative days 2 and 3 compared to those with lower preoperative NLR (11.0 (7.8-16.4) vs. 7.4 (6.2-10.8), P = 0.02; and 11.3 (7.5-17.3) vs. 6.6 (3.3-4.6), P = 0.008, respectively). Despite this, preoperative NLR was not associated with any clinical outcomes. Conclusions Preoperative NLR was not predictive of IVPCA consumption or clinical outcomes following cardiac surgery.

Phenotypic convergence in unrelated lineages arises when different organisms adapt similarly under comparable selective pressures. In an apparent example of this process, syringyl lignin, a fundamental building block of plant cell walls, occurs in two major plant lineages, lycophytes and angiosperms, which diverged from one another more than 400 million years ago. Here, we show that this convergence resulted from independent recruitment of lignin biosynthetic cytochrome P450-dependent monooxygenases that route cell wall monomers through related but distinct pathways in the two lineages. In contrast with angiosperms, in which syringyl lignin biosynthesis requires two phenylpropanoid meta-hydroxylases C3'H and F5H, the lycophyte Selaginella employs one phenylpropanoid dual meta-hydroxylase to bypass several steps of the canonical lignin biosynthetic pathway. Transgenic expression of the Selaginella hydroxylase in Arabidopsis thaliana dramatically reroutes its endogenous lignin biosynthetic pathway, yielding a novel lignin composition not previously identified in nature. Our findings demonstrate a unique case of convergent evolution via distinct biochemical strategies and suggest a new way to genetically reconstruct lignin biosynthesis in higher plants.

Clostridium difficile is the main cause of nosocomial diarrhea, but is also found in asymptomatic subjects that are potentially involved in transmission of C. difficile infection. A sensitive and accurate detection method of C. difficile, especially toxigenic strains is indispensable for the epidemiological investigation. TaqMan-based quantitative-PCR (qPCR) method for targeting 16S rRNA, tcdB, and tcdA genes of C. difficile was developed. The detection limit and accuracy of qPCR were evaluated by analyzing stool samples spiked with known amounts of C. difficile. A total of 235 stool specimens collected from 82 elderly nursing home residents were examined by qPCR, and the validity was evaluated by comparing the detection result with that by C. difficile selective culture (CDSC). The analysis of C. difficile-spiked stools confirmed that qPCR quantified whole C. difficile (TcdA+TcdB+, TcdA-TcdB+, and TcdA-TcdB- types), TcdB-producing strains (TcdA+TcdB+ and TcdA-TcdB+ types), and TcdA-producing strains (TcdA+TcdB+ type), respectively, with a lower detection limit of 103 cells/g of stool. Of the 235 specimens examined, 12 specimens (5.1%) were C. difficile-positive by qPCR: TcdA+TcdB+ strain in six specimens and TcdA-TcdB- strain in the other six. CDSC detected C. difficile in 9 of the 12 specimens, and toxigenic types of the isolates from the 9 specimens were consistent with those identified by qPCR, supporting the validity of our qPCR method. Moreover, the qPCR examination revealed that the carriage rate of whole C. difficile and that of toxigenic strains in the 82 subjects over a 6-month period ranged from 2.4 to 6.8% and 1.2 to 3.8%, respectively. An average qPCR count of C. difficile detected was 104.5 cells/g of stool, suggesting that C. difficile constituted a very small fraction of intestinal microbiota. Our qPCR method should be an effective tool for both clinical diagnosis and epidemiological investigation of C. difficile.