Grab regulates transferrin receptor recycling and iron uptake in developing erythroblasts.

Developing erythroblasts acquire massive amounts of iron through the transferrin (Tf) cycle, which involves endocytosis, sorting, and recycling of the Tf-Tf receptor (Tfrc) complex. Previous studies on the hemoglobin-deficit (hbd) mouse have shown that the exocyst complex is indispensable for the Tfrc recycling; however, the precise mechanism underlying the efficient exocytosis and recycling of Tfrc in erythroblasts remains unclear. Here, we identify the guanine nucleotide exchange factor Grab as a critical regulator of the Tf cycle and iron metabolism during erythropoiesis. Grab is highly expressed in differentiating erythroblasts. Loss of Grab diminishes the Tfrc recycling and iron uptake, leading to hemoglobinization defects in mouse primary erythroblasts, mammalian erythroleukemia cells, and zebrafish embryos. These defects can be alleviated by supplementing iron together with hinokitiol, a small-molecule natural compound that can mediate iron transport independent of the Tf cycle. Mechanistically, Grab regulates the exocytosis of Tfrc-associated vesicles by activating the GTPase Rab8, which subsequently promotes the recruitment of the exocyst complex and vesicle exocytosis. Our results reveal a critical role for Grab in regulating the Tf cycle and provide new insights into iron homeostasis and erythropoiesis.

Trauma Isn't One Size Fits All: How Online Support Communities Point to Different Diagnostic Criteria for C-PTSD and PTSD.

Reddit has provided rich data on mental health discourse. The present study uses 40,335 online posts from Reddit communities to investigate how language can contribute to the understanding of PTSD and C-PTSD. The results showed distinct language patterns in the use of first-person pronouns, cognitive processing, and emotion words, suggesting that they are separate disorders with different effects on survivors. Further, while some social media studies have differentiated submissions and comments, few have investigated the language changes between these contexts. Post-hoc results showed a clear distinction between two contexts across several linguistic markers. Discussion and future directions are explored.

Exosomal miR-222-3p contributes to castration-resistant prostate cancer by activating mTOR signaling.

Despite the clinical benefits of androgen deprivation therapy, most patients with advanced androgen-dependent prostate cancer (ADPC) eventually relapse and progress to lethal androgen-independent prostate cancer (AIPC), also termed castration-resistant prostate cancer (CRPC). MiRNAs can be packaged into exosomes (Exos) and shuttled between cells. However, the roles and mechanisms of exosomal miRNAs involved in CRPC progression have not yet been fully elucidated. Here, we find that miR-222-3p is elevated in AIPC cells, which results in remarkable enhancement of cell proliferation, migration, and invasion ability. Furthermore, Exos released by AIPC cells can be uptaken by ADPC cells, thus acclimating ADPC cells to progressing to more aggressive cell types in vitro and in vivo through exosomal transfer of miR-222-3p. Mechanistically, Exos-miR-222-3p promoted ADPC cells transformed to AIPC-like cells, at least in part, by activating mTOR signaling through targeting MIDN. Our results show that AIPC cells secrete Exos containing miRNA cargo. These cargos can be transferred to ADPC cells through paracrine mechanisms that have a strong impact on cellular functional remodeling. The current work underscores the great therapeutic potential of targeting Exo miRNAs, either as a single agent or combined with androgen receptor pathway inhibitors for CRPC treatment.

Integration of transcriptomics and metabolomics reveals pathways involved in MDSC supernatant attenuation of TGF-ß1-induced myofibroblastic differentiation of mesenchymal stem cells.

Overexposure to transforming growth factor b1 (TGF-ß1) induces myofibroblastic differentiation of mesenchymal stem cells (MSCs), which could be attenuated by myeloid-derived suppressor cell (MDSC) supernatant. However, the promyofibroblastic effects of TGF-ß1 and the antimyofibroblastic effects of MDSC supernatant in MSCs have not been fully elucidated. To further clarify the latent mechanism and identify underlying therapeutic targets, we used an integrative strategy combining transcriptomics and metabolomics. Bone marrow MSCs were collected 24 h following TGF-ß1 and MDSC supernatant treatment for RNA sequencing and untargeted metabolomic analysis. The integrated data were then analyzed to identify significant gene-metabolite correlations. Differentially expressed genes (DEGs) and differentially expressed metabolites (DEMs) were assessed by Gene Ontology (GO) functional annotation and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway analyses for exploring the mechanisms of myofibroblastic differentiation of MSCs. The integration of transcriptomic and metabolomic data highlighted significantly coordinated changes in glycolysis/gluconeogenesis and purine metabolism following TGF-ß1 and MDSC supernatant treatment. By combining transcriptomic and metabolomic analyses, this study showed that glycolysis/gluconeogenesis and purine metabolism were essential for the myofibroblastic differentiation of MSCs and may serve as promising targets for mechanistic research and clinical practice in the treatment of fibrosis by MDSC supernatant.

A diagrammatic method for the identification and resolution of urban spatial conflicts.

Spatial conflicts are formed in the process of urbanization and become the primary drivers of urban ecological and environmental problems. The defining trait of a spatial conflict is the occupation of ecological or agricultural spaces by construction spaces. This work presents a classification scheme for spatial conflicts in China, including source-area conflicts, corridor conflicts, safety conflicts, and farmland conflicts. Also, it constructs a model for the determination of spatial conflicts and conflict intensity using diagrammatic method. Based on a case study performed using our methods on Shenzhen, we proposed a timeline and policy roadmap for the resolution of spatial conflicts in Shenzhen according to the severity and characteristics of spatial conflicts in the city. The results show that the total spatially conflicted area of Shenzhen is 10.57 km2, and the percentage of construction land-use in these areas is 1.37%. The spatial conflicts are mainly source-area or corridor conflicts, and minor conflicts account for approximately 60% of the total conflicted area. Most of the spatial conflicts are either "easy to resolve" or "moderately easy to resolve". Overall, in terms of the severity of spatial conflict, Shenzhen remains at the "stable and under control" level. Considering the primary aim of spatial conflict resolution is to revert built-up lands into urban green spaces, we proposed a timeline for the resolution of spatial conflicts in Shenzhen over the next 15 years, as well as a system of supporting policies. The results of this study shall serve as a guide for the optimization of urban spatial structures and the promotion of sustainable urban development.

Pediatric Consultation-Liaison: Patient Characteristics and Considerations for Training in Evidence-Based Practices.

Consultation-liaison services are an integral part of many pediatric hospital settings, yet characteristics of this patient population have not been extensively documented. The current study is a retrospective one-year chart review of the consultation-liaison service at a large pediatric hospital in the Southwestern United States. The purpose of this study is twofold: (1) to characterize this hospital's CL population and (2) to use these characteristics to identify preliminary evidence-based practices that should be considered for CL provider training. Identifying evidence-based practice elements that align with the characteristics of consultation-liaison patient populations may inform trainings for consultation-liaison staff. This would help to ensure that youth seen in hospital consultation-liaison services are getting the best available services, which is critical given the shortened time frame available to work with this patient population.

Fast and Automatic Reconstruction of Semantically Rich 3D Indoor Maps from Low-quality RGB-D Sequences.

Semantically rich indoor models are increasingly used throughout a facility's life cycle for different applications. With the decreasing price of 3D sensors, it is convenient to acquire point cloud data from consumer-level scanners. However, most existing methods in 3D indoor reconstruction from point clouds involve a tedious manual or interactive process due to line-of-sight occlusions and complex space structures. Using the multiple types of data obtained by RGB-D devices, this paper proposes a fast and automatic method for reconstructing semantically rich indoor 3D building models from low-quality RGB-D sequences. Our method is capable of identifying and modelling the main structural components of indoor environments such as space, wall, floor, ceilings, windows, and doors from the RGB-D datasets. The method includes space division and extraction, opening extraction, and global optimization. For space division and extraction, rather than distinguishing room spaces based on the detected wall planes, we interactively define the start-stop position for each functional space (e.g., room, corridor, kitchen) during scanning. Then, an interior elements filtering algorithm is proposed for wall component extraction and a boundary generation algorithm is used for space layout determination. For opening extraction, we propose a new noise robustness method based on the properties of convex hull, octrees structure, Euclidean clusters and the camera trajectory for opening generation, which is inapplicable to the data collected in the indoor environments due to inevitable occlusion. A global optimization approach for planes is designed to eliminate the inconsistency of planes sharing the same global plane, and maintain plausible connectivity between the walls and the relationships between the walls and openings. The final model is stored according to the CityGML3.0 standard. Our approach allows for the robust generation of semantically rich 3D indoor models and has strong applicability and reconstruction power for complex real-world datasets.

Tin dioxide nanoparticles as catalyst precursors for plasma-assisted vapor-liquid-solid growth of silicon nanowires with well-controlled density.

The fabrication of arrays of silicon nanowires (Si NWs) with well-defined surface coverage using the vapor-liquid-solid process requires a good control of the density and size distribution for the metal catalyst. We report on a cost-effective bottom-up approach to produce Si NWs by a low-temperature deposition technology using plasma-enhanced chemical vapor deposition and tin dioxide (SnO2) nanoparticles as the source of tin catalyst. This strategy offers a straightforward method to select specific particle sizes by conventional colloidal techniques, and to tune the surface coverage using a polyelectrolyte layer to efficiently immobilize the particles on the substrate by electrostatic grafting. After a further step of reduction into tin metal droplets using hydrogen plasma treatment, the catalyst particles are used for the growth of Si NWs. This approach allows the prodcution of controlled Si NWs arrays which can be used as a template for radial junction thin film solar cells.

Geometric Integration of Hybrid Correspondences for RGB-D Unidirectional Tracking.

Traditionally, visual-based RGB-D SLAM systems only use correspondences with valid depth values for camera tracking, thus ignoring the regions without 3D information. Due to the strict limitation on measurement distance and view angle, such systems adopt only short-range constraints which may introduce larger drift errors during long-distance unidirectional tracking. In this paper, we propose a novel geometric integration method that makes use of both 2D and 3D correspondences for RGB-D tracking. Our method handles the problem by exploring visual features both when depth information is available and when it is unknown. The system comprises two parts: coarse pose tracking with 3D correspondences, and geometric integration with hybrid correspondences. First, the coarse pose tracking generates the initial camera pose using 3D correspondences with frame-by-frame registration. The initial camera poses are then used as inputs for the geometric integration model, along with 3D correspondences, 2D-3D correspondences and 2D correspondences identified from frame pairs. The initial 3D location of the correspondence is determined in two ways, from depth image and by using the initial poses to triangulate. The model improves the camera poses and decreases drift error during long-distance RGB-D tracking iteratively. Experiments were conducted using data sequences collected by commercial Structure Sensors. The results verify that the geometric integration of hybrid correspondences effectively decreases the drift error and improves mapping accuracy. Furthermore, the model enables a comparative and synergistic use of datasets, including both 2D and 3D features.

Clinical features and prognosis of thymic neuroendocrine tumours associated with multiple endocrine neoplasia type 1: A single-centre study, systematic review and meta-analysis.

OBJECTIVE: Thymic neuroendocrine tumour (TH-NET) accounts for almost 20% of multiple endocrine neoplasia type 1 (MEN1)-associated mortality. Identifying risk factors for the development of these rare tumours and prognostic factors for clinical outcomes will be helpful in clinical practice. DESIGN AND PATIENTS: We performed a retrospective analysis of patients treated for TH-NET associated with MEN1 in a single institution and meta-analysis of literature reports. We used a fixed effect model to pool results across studies to evaluate the prevalence, clinical features and prognosis. RESULTS: TH-NET was detected in 9 (7.4%) of 121 patients with MEN1 seen in our institution, and 5 (55.6%) were women. Seven additional studies were identified through a systematic review of the literature. The pool estimate of TH-NET prevalence was 3.7% (n = 99) in MEN1 (n = 2710), sex ratio was 79:20 (male vs female), and the median age at diagnosis was 43.0 years (range, 16.0-72.0 years). Forty-three patients died with a median survival time of 8.4 years. Older age at diagnosis (HR = 1.4, 95% CI = 1.0-1.8, P = .03), maximum tumour diameter (HR = 1.5, 95% CI = 1.0-2.3, P = .04) and presence of metastasis (HR = 1.6, 95% CI = 1.0-2.5, P = .04) were associated with worse outcome. A male predominance (91.9% vs 59.5%, P < .001) and history of smoking (59.0% vs 23.5%, P = .015) were more common in American/European series compared to Asian reports. CONCLUSION: TH-NET is a rare but fatal component of MEN1. Earlier detection of TH-NET in patients with MEN1 may be recommended which should theoretically result in better outcomes. Different genetic backgrounds (race) appear to result in clinical difference.

The ras-GTPase activity of neurofibromin restrains ERK-dependent FGFR signaling during endochondral bone formation.

The severe defects in growth plate development caused by chondrocyte extracellular signal-regulated protein kinases 1 and 2 (ERK1/2) gain or loss-of-function suggest that tight spatial and temporal regulation of mitogen-activated protein kinase signaling is necessary to achieve harmonious growth plate elongation and structure. We provide here evidence that neurofibromin, via its Ras guanosine triphosphatase -activating activity, controls ERK1/2-dependent fibroblast growth factor receptor (FGFR) signaling in chondrocytes. We show first that neurofibromin is expressed in FGFR-positive prehypertrophic and hypertrophic chondrocytes during growth plate endochondral ossification. Using mice lacking neurofibromin 1 (Nf1) in type II collagen-expressing cells, (Nf1col2(-/-) mutant mice), we then show that lack of neurofibromin in post-mitotic chondrocytes triggers a number of phenotypes reminiscent of the ones observed in mice characterized by FGFR gain-of-function mutations. Those include dwarfism, constitutive ERK1/2 activation, strongly reduced Ihh expression and decreased chondrocyte proliferation and maturation, increased chondrocytic expression of Rankl, matrix metalloproteinase 9 (Mmp9) and Mmp13 and enhanced growth plate osteoclastogenesis, as well as increased sensitivity to caspase-9 mediated apoptosis. Using wildtype (WT) and Nf1(-/-) chondrocyte cultures in vitro, we show that FGF2 pulse-stimulation triggers rapid ERK1/2 phosphorylation in both genotypes, but that return to the basal level is delayed in Nf1(-/-) chondrocytes. Importantly, in vivo ERK1/2 inhibition by daily injection of a recombinant form of C-type natriuretic peptide to post-natal pups for 18 days was able to correct the short stature of Nf1col2(-/-) mice. Together, these results underscore the requirement of neurofibromin and ERK1/2 for normal endochondral bone formation and support the notion that neurofibromin, by restraining RAS-ERK1/2 signaling, is a negative regulator of FGFR signaling in differentiating chondrocytes.

Alterations of Group I mGluRs and BDNF Associated with Behavioral Abnormity in Prenatally Stressed Offspring Rats.

Substantial evidence reveals that prenatal stress is closely linked with abnormal behavior in offspring, but the mechanism remains unclear. In this study, our aim was to observe the alterations of behaviors, metabotropic glutamate receptor-1/5 (mGluR1/5) and brain-derived neurotrophic factor (BDNF) in various brain regions of prenatally stressed offspring rats. The forced swimming test (FST) and the open field test (OFT) were carried on 1-month-old offspring rats. The expression levels of mGluR1, mGluR5, and BDNF mRNA were measured in various brain regions of the offspring rats. Our results showed that the immobile time in the FST was significantly increased in the late prenatal stress (LPS) group compared with that in the control group, especially in the female rats. Similarly, in the OFT, the rats in both the mid prenatal stress (MPS) and LPS groups demonstrated anxiety-like behavior, especially the male rats in the LPS group. The expression of mGluR1 protein was increased in the hippocampus and prefrontal cortex of rats from the LPS group, as well as in the prefrontal cortex of rats from the MPS group. Meanwhile, the expression of mGluR5 protein was facilitated in the hippocampus and prefrontal cortex of rats in the LPS group. The expression of mGluR5 protein was increased in the striatum of the female rats in both MPS and LPS groups, and only in the male rats from the LPS group. In addition, the reduced BDNF mRNA level was detected in the hippocampus and prefrontal cortex in the LPS rats, and in the striatum of the female rats in LPS group. These results indicate that alterations of the mGluR1, mGluR5 and BDNF mRNA may contribute to the depression-like and anxiety-like behaviors of prenatally stressed offspring rats.

[Optimization of trypsin digestion intensity to obtain high-purity in vitro cultured astrocytes].

The aim of the present study was to observe the effect of trypsin digestion on the purity of in vitro cultured astrocytes and optimize the culture methods. The cerebral cortical tissue from newborn Sprague Dawley (SD) rats was isolated and digested with 0.25% trypsin for 20, 30, or 40 min. The obtained single cell suspension was then cultured. Once reaching confluence, the cells were shaken at a constant temperature. Then, each of 20 and 30 min groups was subdivided into two groups, the control group with normal digestion and two-time-digestion group, and the cells were passaged and purified. Through inverted phase contrast microscope and MTT assay, cell growth and proliferation were observed, respectively. Immunofluorescence for glial fibrillary acidic protein (GFAP) was used to observe the morphology of astrocytes and to assess their purity in different stages. Flow cytometric analysis was used to detect the apoptotic rates of purified astrocytes. The results showed that, the cells being digested for 20 min usually reached confluence at 9 d after seeding. When the digestion time was extended to 30 min, the cells grew faster and reached confluence at 7 d after seeding, meanwhile the morphology of astrocytes was normal, GFAP positive rate (70.2 ± 4.0)% being much higher than that of the 20 min group (P < 0.05). Compared with 20 min group, 40 min group showed higher GFAP positive rate, whereas the cell proliferation was slower, and cell injury was more obvious. After shaking at constant temperature, two times of trypsin digestion could decrease the number of contaminated cells after passage. The GFAP positive rates of two-time-digestion groups in passage 1 (P1) were higher than those of corresponding control groups, and the GFAP positive rate of 30 min + two-time-digestion group in P1 reached (98.1 ± 1.7)%, which was equivalent to that of the 20 min + control group in P3. However, the apoptotic rate showed no significant difference between these two groups. Based on above mentioned results, we conclude that 30 min + two-time of trypsin digestion effectively improves the purity of astrocytes and shortens the time of primary culture and purification, suggesting that it is a rapid and effective method to obtain astrocytes with high purity in vitro.

Taurine attenuates amyloid ß 1-42-induced mitochondrial dysfunction by activating of SIRT1 in SK-N-SH cells.

Amyloid ß (Aß) plays a critical role in the pathogenesis of Alzheimer disease (AD). Studies indicate that Aß causes reactive oxygen species (ROS) generation, mitochondrial dysfunction and neurons loss in vivo and in vitro. Taurine, a naturally occurring ß-amino acid in the brain, has been demonstrated to have neuroprotective properties. In the present study, the effects of taurine on cell viability and mitochondrial function in Aß1-42-treated SK-N-SH cells were investigated. Pretreatment of taurine significantly attenuated Aß1-42-induced neuronal death. Similarly, taurine suppressed the mPTP opening and reversed mitochondrial function in the presence of Aß1-42. Additionally, taurine attenuated the intracellular Ca(2+) and ROS generation induced by Aß1-42. Moreover, the expression of Sirtuin 1 (SIRT1) was obviously recovered by taurine in Aß1-42-treated SK-N-SH cells. Our results suggest that taurine prevents Aß1-42-induced mitochondrial dysfunction by activation of SIRT1. This study implies that taurine is a prospective additive for AD patients.

Activation of SIRT1 by curcumin blocks the neurotoxicity of amyloid-ß25-35 in rat cortical neurons.

As one of the most important hallmarks of Alzheimer's disease (AD), ß-amyloid (Aß) plays important roles in inducing reactive oxygen species (ROS) generation, mitochondrial dysfunction and apoptotic cell death in neurons. Curcumin extracted from the yellow pigments spice plant turmeric shows multiplied bioactivities such as antioxidant and anti-apoptosis properties in vitro and in vivo. In the present study, the neuroprotective effect of curcumin against Aß25-35-induced cell death in cultured cortical neurons was investigated. We found that pretreatment of curcumin prevented the cultured cortical neurons from Aß25-35-induced cell toxicity. In addition, curcumin improved mitochondrial membrane potential (ΔΨm), decreased ROS generation and inhibited apoptotic cell death in Aß25-35 treated neurons. Furthermore, we found that application of curcumin activated the expression of SIRT1 and subsequently decreased the expression of Bax in the presence of Aß25-35. The protective effect of curcumin was blocked by SIRT1 siRNA. Taken together, our results suggest that activation of SIRT1 is involved in the neuroprotective action of curcumin.

Identification of genes and key pathways underlying the pathophysiological association between sarcopenia and chronic obstructive pulmonary disease.

PURPOSE: Chronic obstructive pulmonary disease (COPD) patients are likely to develop sarcopenia, while the exact mechanism underlying the association between sarcopenia and COPD is still not clear. This cohort study aims to explore the genes, signaling pathways, and transcription factors (TFs) that are related to the molecular pathogenesis of sarcopenia and COPD. METHODS: According to the strict inclusion criteria, two gene sets (GSE8479 for sarcopenia and GSE76925 for COPD) were obtained from the Gene Expression Omnibus (GEO) platform. Overlapping differentially expressed genes (DEGs) in sarcopenia and COPD were detected, and comprehensive bioinformatics analysis was conducted, including functional annotation, enrichment analysis of Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG), construction of a protein-protein interaction (PPI) network, co-expression analysis, identification and validation of hub genes, and TFs prediction and verification. RESULTS: In total, 118 downregulated and 92 upregulated common DEGs were detected. Functional analysis revealed that potential pathogenesis involves oxidoreductase activity and ferroptosis. Thirty hub genes were detected, and ATP metabolic process and oxidative phosphorylation were identified to be closely related to the hub genes. Validation analysis revealed that SAA1, C3, and ACSS2 were significantly upregulated, whereas ATF4, PPARGC1A, and MCTS1 were markedly downregulated in both sarcopenia and COPD. In addition, six TFs (NFKB1, RELA, IRF7, SP1, MYC, and JUN) were identified to regulate the expression of these genes, and SAA1 was found to be coregulated by NFKB1 and RELA. CONCLUSION: This study uncovers potential common mechanisms of COPD complicated by sarcopenia. The hub gene SAA1 and the NF-κB signaling pathway could be involved, and oxidative phosphorylation and ferroptosis might be important contributors to this comorbidity.

Analysis of railway accessibility in Fujian Province and the influence of economic development on its spatial differentiation.

Understanding railway accessibility supports railway regulation and development, but few studies consider different perspectives. We study the spatial distribution of accessibility indicators at the county (city) scale in Fujian Province by spatiotemporal syntax and weighted average travel time using railway timetable data. Export trade, rich commercial activities, and high-speed rail had a significant positive effect on objective accessibility. Fuzhou, Sanming, and Longyan were main transfer centers. The most accessible nodes based on weighted average travel time formed a "U"-shaped corridor along the coast. The county-wide average accessibility was 1.72 h. According to spatiotemporal syntax, local general public budget expenditure (0.758993) and export volume of goods-total retail sales of consumer goods (0.956257) had the most interactive impact, while according to weighted average accessibility, import volume of goods (0.618447) and per capita gross regional product-import volume of goods (0.878573) did. These results provide reference for transportation planning and regional development in Fujian Province.

Tapping into alcohol use during COVID: Drinking correlates among bartenders and servers.

The COVID pandemic placed a spotlight on alcohol use and the hardships of working within the food and beverage industry, with millions left jobless. Following previous studies that have found elevated rates of alcohol problems among bartenders and servers, here we studied the alcohol use of bartenders and servers who were employed during COVID. From February 12-June 16, 2021, in the midst of the U.S. COVID national emergency declaration, survey data from 1,010 employed bartender and servers were analyzed to quantify rates of excessive or hazardous drinking along with regression predictors of alcohol use as assessed by the 10-item Alcohol Use Disorders Identification Test (AUDIT). Findings indicate that more than 2 out of 5 (44%) people surveyed reported moderate or high rates of alcohol problem severity (i.e., AUDIT scores of 8 or higher)-a rate 4 to 6 times that of the heavy alcohol use rate reported pre- or mid-pandemic by adults within and outside the industry. Person-level factors (gender, substance use, mood) along with the drinking habits of one's core social group were significantly associated with alcohol use. Bartenders and servers reported surprisingly high rates of alcohol problem severity and experienced risk factors for hazardous drinking at multiple ecological levels. Being a highly vulnerable and understudied population, more studies on bartenders and servers are needed to assess and manage the true toll of alcohol consumption for industry employees.

Mice lacking Nf1 in osteochondroprogenitor cells display skeletal dysplasia similar to patients with neurofibromatosis type I.

Mutations in NF1 cause neurofibromatosis type I (NF1), a disorder characterized, among other clinical manifestations, by generalized and focal bony lesions. Dystrophic scoliosis and tibial pseudoarthrosis are the most severe skeletal manifestations for which treatment is not satisfactory, emphasizing the dearth of knowledge related to the biology of NF1 in bone cells. Using reporter mice, we report here that the mouse Col2α1-Cre promoter (collagen, type II, alpha 1) is active not only in chondrocytes but also in adult bone marrow osteoprogenitors giving rise to osteoblasts. Based on this finding, we crossed the Col2α1-Cre transgenic and Nf1(flox/flox) mice to determine whether loss of Nf1 in axial and appendicular osteochondroprogenitors recapitulates the skeletal abnormalities of NF1 patients. By microtomographic and X-rays studies, we show that Nf1(Col2)(-/-) mice display progressive scoliosis and kyphosis, tibial bowing and abnormalities in skull and anterior chest wall formation. These defects were accompanied by a low bone mass phenotype, high bone cortical porosity, osteoidosis, increased osteoclastogenesis and decreased osteoblast number, as quantified by histomorphometry and 3D-microtomography. Loss of Nf1 in osteochondroprogenitors also caused severe short stature and intervertebral disc defects. Blockade of the RAS/ERK activation characteristic of Nf1(-/-) osteoprogenitors by lovastatin during embryonic development could attenuate the increased cortical porosity observed in mutant pups. These data and the skeletal similarities between this mouse model and NF1 patients thus suggest that activation of the RAS/ERK pathway by Nf1 loss-of-function in osteochondroprogenitors is responsible for the vertebral and tibia lesions in NF1 patients, and that this molecular signature may represent a good therapeutic target.

[Synergistic action of 4-hydroxynonenal and tumor necrosis factor involving the NF-kB/IkBa signaling pathway in alcohol-induced liver injury].

OBJECTIVE: To investigate the effects and mechanism of intracellular 4-hydroxynonenal (4-HNE) accumulation on tumor necrosis factor (TNF)-induced hepatotoxicity in alcoholic liver disease (ALD). METHODS: An ALD model was established in male C57BL/6 mice (6-8 weeks old) by feeding an ethanol-containing diet for 5 weeks; mice given regular (non-ethanol) diet served as controls. ALD-related changes in 4-HNE and TNF levels were detected by western blotting. The underlying mechanisms of this molecular effect were examined by pre-treating HepG2 cells with 4-HNE followed by exposure to various concentrations of TNF. Effects on cell death were evaluated by MTT assay. Effects on TNF-mediated upstream factors' expression were detected by ELISA, western blotting, and real-time PCR. Effects on the TNF-induced inhibitor of NF-kB (IkBa) activity (phosphorylation status) and its formation of adducts were detected by western blotting and immunoprecipitation. RESULTS: ALD mice showed increased hepatic 4-HNE and TNF levels, and the increases were associated with extent of liver injury. Cell culture studies revealed that 4-HNE, at non-toxic concentrations, sensitized hepatocytes to TNF killing, which was associated with suppressed NF-kB trans activity. Furthermore, 4-HNE prevented phosphorylation of IkBa without affecting upstream IkB kinase activity. The ALD-enhanced 4-HNE content was found to associated with increased formation of 4-HNE-IkBa adduction for both the 4-HNE - treated hepatocytes in culture and in the livers of ALD mice. CONCLUSION: Alcohol-induced increase in 4-HNE accumulation represents a potent and clinically relevant mechanism of sensitizing hepatocytes to TNF-induced toxicity. These data support the notion that decreasing or eliminating accumulated intracellular 4-HNE can serve as a potential therapeutic option for ALD.