Direct Construction of 1,4-Dihydropyridazines and Pyrazoles via Annulation of Alkyl 2-Aroyl-1-chlorocyclopropanecarboxylates.

An efficient and chemodivergent synthesis of highly functionalized 1,4-dihydropyridazines and pyrazoles has been accomplished via base-promoted annulation between hydrazones and alkyl 2-aroyl-1-chlorocyclopropanecarboxylates, respectively. This transition-metal-free domino reaction proceeded rapidly under mild basic conditions, affording potentially bioactive 1,4-dihydropyridazine and pyrazole derivatives in moderate yields. The conversion of 1,4-dihydropyridazine to pyrazole was confirmed by adjusting the quantity of the base.

[Prenatal diagnosis and genetic analysis of three fetuses with paternal chromosomal simplex 3q microduplication syndrome].

OBJECTIVE: To explore the phenotypic characteristics of paternal chromosomal simplex 3q microduplication syndrome. METHODS: Amniotic fluid samples of 3 fetuses from a same couple were subjected to prenatal diagnosis through combined high-resolution chromosomal G-banding karyotyping and chromosomal microarray analysis (CMA). Peripheral blood samples were also collected the couple for the determination of parental origin. RESULTS: The karyotypes of all three fetuses were 46,XN,dup(3)(q25q26.1), and their CMA results were arr[hg19]3q25.33q26.1(159 336 333-166 924 969)×3. The duplication in the three fetuses have all derived from their father. No anomaly with found with the mother by CMA . CONCLUSION: Through combined G-banded chromosomal karyotyping and CMA assay, a paternally derived 3q25.33-q26.1 microduplication has been identified, which has enabled genetic counseling for this couple.

Increased regional homogeneity modulated by metacognitive training predicts therapeutic efficacy in patients with schizophrenia.

Previous studies have demonstrated the efficacy of metacognitive training (MCT) in schizophrenia. However, the underlying mechanisms related to therapeutic effect of MCT remain unknown. The present study explored the treatment effects of MCT on brain regional neural activity using regional homogeneity (ReHo) and whether these regions' activities could predict individual treatment response in schizophrenia. Forty-one patients with schizophrenia and 20 healthy controls were scanned using resting-state functional magnetic resonance imaging. Patients were randomly divided into drug therapy (DT) and drug plus psychotherapy (DPP) groups. The DT group received only olanzapine treatment, whereas the DPP group received olanzapine and MCT for 8 weeks. The results revealed that ReHo in the right precuneus, left superior medial prefrontal cortex (MPFC), right parahippocampal gyrus and left rectus was significantly increased in the DPP group after 8 weeks of treatment. Patients in the DT group showed significantly increased ReHo in the left ventral MPFC/anterior cingulate cortex (ACC), left superior MPFC/middle frontal gyrus (MFG), left precuneus, right rectus and left MFG, and significantly decreased ReHo in the bilateral cerebellum VIII and left inferior occipital gyrus (IOG) after treatment. Support vector regression analyses showed that high ReHo levels at baseline in the right precuneus and left superior MPFC could predict symptomatic improvement of Positive and Negative Syndrome Scale (PANSS) after 8 weeks of DPP treatment. Moreover, high ReHo levels at baseline and alterations of ReHo in the left ventral MPFC/ACC could predict symptomatic improvement of PANSS after 8 weeks of DT treatment. This study suggests that MCT is associated with the modulation of ReHo in schizophrenia. ReHo in the right precuneus and left superior MPFC may predict individual therapeutic response for MCT in patients with schizophrenia.

Increased Homotopic Connectivity in the Prefrontal Cortex Modulated by Olanzapine Predicts Therapeutic Efficacy in Patients with Schizophrenia.

Background: Previous studies have revealed the abnormalities in homotopic connectivity in schizophrenia. However, the relationship of these deficits to antipsychotic treatment in schizophrenia remains unclear. This study explored the effects of antipsychotic therapy on brain homotopic connectivity and whether the homotopic connectivity of these regions might predict individual treatment response in schizophrenic patients. Methods: A total of 21 schizophrenic patients and 20 healthy controls were scanned by the resting-state functional magnetic resonance imaging. The patients received olanzapine treatment and were scanned at two time points. Voxel-mirrored homotopic connectivity (VMHC) and pattern classification techniques were applied to analyze the imaging data. Results: Schizophrenic patients presented significantly decreased VMHC in the temporal and inferior frontal gyri, medial prefrontal cortex (MPFC), and motor and low-level sensory processing regions (including the fusiform gyrus and cerebellum lobule VI) relative to healthy controls. The VMHC in the superior/middle MPFC was significantly increased in the patients after eight weeks of treatment. Support vector regression (SVR) analyses revealed that VMHC in the superior/middle MPFC at baseline can predict the symptomatic improvement of the positive and negative syndrome scale after eight weeks of treatment. Conclusions: This study demonstrated that olanzapine treatment may normalize decreased homotopic connectivity in the superior/middle MPFC in schizophrenic patients. The VMHC in the superior/middle MPFC may predict individual response for antipsychotic therapy. The findings of this study conduce to the comprehension of the therapy effects of antipsychotic medications on homotopic connectivity in schizophrenia.

Comparative analysis of mesenchymal stromal cells derived from rabbit bone marrow and Wharton's jelly for adipose tissue engineering.

Purpose: To investigate the proliferative, adipogenic, and immunological properties of rabbit Mesenchymal stromal cells (MSCs) derived from bone marrow and umbilical cord Wharton's jelly.Materials and Methods: We extracted rabbit MSCs from bone marrow (BMSCs) and umbilical cord Wharton's jelly (WJ-MSCs). Both BMSCs and WJ-MSCs underwent adipogenic differentiation for 2 weeks, and then were transferred to non-inductive complete medium. Their adipogenic capacities were examined by histomorphometry and quantitative RT-PCR (qRT-PCR). The immunological markers were determined by mRNA expression of MHC-Ia, MHC-II, and RLA-DRA by qRT-PCR and protein expression of MHC-II by immunofluorescent staining. The proliferative capacities of adipogenic MSCs were also examined by counting kit-8 experiment and cell population doubling time.Results: We found that adipogenic differentiation increased the mRNA expression levels of adipogenic and immunological markers. The protein expression levels of MHC-II also increased after adipogenic differentiation in both groups. The adipogenic BMSCs showed higher mRNA expression levels of adipogenic and immunological markers. Removal of adipogenic agents after 2 weeks of adipo-differentiation inversely decreased the expression of immunological and adipogenic markers. The adipo-differentiation could decreased the proliferative capacities of both MSCs, but the adipogenic WJ-MSCs showed significantly higher proliferative capacities than BMSCs.Conclusions: Adipogenic differentiation increased the immunogenicity of both BMSCs and WJ-MSCs, and dedifferentiation inversely decreased their immunogenicity. Adipogenic WJ-MSCs showed significantly higher proliferative and immunoprivileged capacities than BMSCs, and the dedifferentiated BMSCs showed almost the same adipogenic capacity as WJ-MSCs. WJ-MSCs were more suitable than BMSCs for adipose tissue engineering.

Synthetic Entry to the 2-Azatricyclo[4.3.2.04,9]undecane Ring System via Tropone.

A synthesis of the 2-azatricyclo[4.3.2.04,9]undecane ring system-a hitherto unreported bridged azatricyclic ring system-beginning from tricarbonyl(tropone)iron and allylamine was accomplished in three steps: (1) aza-Michael addition of allylamine to tricarbonyl(tropone)iron; (2) Boc-protection of the resulting secondary amine; and (3) oxidative demetallation leading to a spontaneous intramolecular Diels-Alder reaction. The effect of a variety of parameters on the intramolecular Diels-Alder reaction was investigated, including diene and dienophile substitution patterns and dienophile tether length.

Exosomal MMP2 derived from mature osteoblasts promotes angiogenesis of endothelial cells via VEGF/Erk1/2 signaling pathway.

The skeletal system is a dynamic organ that continuously undergoes coupled trabeculae and blood vessels remodeling, indicating the possible existence of molecular crosstalk between endothelial and osteoblastic cells. Since the cross-talk between bone-forming osteoblasts (OBs) and vessel-forming endothelial cells (ECs) have progressively gained investigators' attention, few studies focused on the regulatory function of extracellular vesicles derived from OBs on ECs. In this study, the effect of the exosomes derived from mature osteoblasts (MOBs) on the ECs was investigated. Firstly, exosomes derived from mature osteoblasts (MOB-Exos) were isolated and identified by NanoSight light scatter technology, electron microscopy and Western bolting. Fluorescent labeling of MOB-Exos revealed its internalization by ECs. RNA interference technique was used to knock down matrix metalloproteinase-2 (MMP2) in MOB-Exos. Then ECs were co-cultured with MOB-Exos and MMP2 knockdown MOB-Exos. Wound healing migration assay, transwell migration assay, CCK-8 assay and tube formation assay of ECs were conducted to determine the angiogenic capability of ECs. Then the VEGF/Erk1/2 pathway markers were detected by Western blot. Our results showed that MOB-Exos could promote the proliferation, migration and tube formation of ECs. Meanwhile, the promoted angiogenetic capacities of ECs were impaired when MMP2 in MOB-Exos was knocked down. In addition, immunoblotting indicated that MOB-Exos could promote the activation of the VEGF/Erk1/2 pathway of ECs; whereas the activation of the VEGF/Erk1/2 pathway was attenuated when the ECs were co-cultured with the MMP2 knockdown MOB-Exos. In conclusion, the MMP-2 existing in exosomes derived from MOBs could promote the angiogenesis of ECs in vitro, which might be realized through VEGF/Erk1/2 signaling pathway.

Incorporating Negative Sample Training for Ship Detection Based on Deep Learning.

While ship detection using high-resolution optical satellite images plays an important role in various civilian fields-including maritime traffic survey and maritime rescue-it is a difficult task due to influences of the complex background, especially when ships are near to land. In current literatures, land masking is generally required before ship detection to avoid many false alarms on land. However, sea⁻land segmentation not only has the risk of segmentation errors, but also requires expertise to adjust parameters. In this study, Faster Region-based Convolutional Neural Network (Faster R-CNN) is applied to detect ships without the need for land masking. We propose an effective training strategy for the Faster R-CNN by incorporating a large number of images containing only terrestrial regions as negative samples without any manual marking, which is different from the selection of negative samples by targeted way in other detection methods. The experiments using Gaofen-1 satellite (GF-1), Gaofen-2 satellite (GF-2), and Jilin-1 satellite (JL-1) images as testing datasets under different ship detection conditions were carried out to evaluate the effectiveness of the proposed strategy in the avoidance of false alarms on land. The results show that the method incorporating negative sample training can largely reduce false alarms in terrestrial areas, and is superior in detection performance, algorithm complexity, and time consumption. Compared with the method based on sea⁻land segmentation, the proposed method achieves the absolute increment of 70% of the F1-measure, when the image contains large land area such as the GF-1 image, and achieves the absolute increment of 42.5% for images with complex harbors and many coastal ships, such as the JL-1 images.

[The mitochondrial tRNAMet/tRNAGlnA4401G and tRNACysG5821A mutations may be associated with hypertension in two Han Chinese families].

Mitochondrial tRNA genes are the hot spots for mutations associated with essential hypertension. We report here the clinical and molecular genetic characterization of two Han Chinese pedigrees with materially inherited essential hypertension. Clinical evaluation revealed the variable severity and age-at-onset of hypertension among matrilineal relatives. In particular, the age-at-onset of hypertension in the maternal kindred ranged from 36 years to 79 years. The sequence analysis of entire mitochondrial genome in two probands showed that two probands carried the identical homoplasmic tRNAMet/tRNAGlnA4401G and tRNACysG5821A mutations and distinct sets of polymorphisms belonging to East Asian haplogroup C. The A4401G mutation may affect the processing of the precursors of tRNAMet and tRNAGln , thereby altering the tRNA metabolism. The tRNACys G5821A mutation is located in the acceptor stem of tRNACys. This mutation may abol-ish the predicted G6-C67 pairing and consequently affect the structure and stability of mitochondrial tRNACys, thereby leading to mitochondrial dysfunction. Therefore, these data suggested that the tRNAMet/tRNAGlnA4401G and tRNACys G5821A mutations are likely associated with essential hypertension in these two Chinese pedigrees.

Exosomal miR-423-5p Derived from Cerebrospinal Fluid Pulsation Stress-Stimulated Osteoblasts Improves Angiogenesis of Endothelial Cells via DUSP8/ERK1/2 Signaling Pathway.

Exosomes secreted from osteoblasts (OBs) can regulate the angiogenesis of endothelial cells (ECs); however, whether cerebrospinal fluid pulsation (CSFP) stress, a special mechanical stimulation, can influence the cell's communication in the context of angiogenesis remains unknown. In this study, the effect of exosomes derived from CSFP stress-stimulated OBs on facilitating the angiogenesis of ECs was investigated. First, OBs were cultured in a CSFP bioreactor, and exosomes derived from OBs were isolated and identified. Cell Counting Kit 8 assay, transwell migration assay, wound healing migration assay, and tube formation assay were conducted to assess the effects of CSFP stress-stimulated OBs-derived exosomes (CSFP-Exos) on the angiogenesis of ECs. Then high-throughput RNA sequencing was used to determine the miRNA profiles of Non-CSFP stress-stimulated OBs-derived exosomes (NCSFP-Exos) and CSFP-Exos, and the luciferase reporter gene assay was performed to confirm the binging of miR-423-5p to DUSP8. In addition, the Matrigel plug assay was performed to explore whether exosomal miR-423-5p has the same effects in vivo. Our results suggested that CSFP-Exos can promote the angiogenesis of ECs, and miR-423-5p was enriched in CSFP-Exos. Moreover, miR-423-5p could promote the effect of angiogenesis via directly targeting dual-specificity phosphatase 8 (DUSP8), which inhibited the ERK1/2 signaling pathway. In conclusion, exosomal miR-423-5p derived from CSFP stress-stimulated OBs could promote the angiogenesis of ECs by the DUSP8/ERK1/2 signaling pathway.

[Research progress in heritable dyslipidemia].

Dyslipidemia is defined as high levels of serum cholesterol and/or triglycerides. Dyslipidemia often leads to severe cardiovascular diseases including coronary heart disease and stroke as the first clinical manifestation, thus threatening the health of human beings. Dyslipidemia diseases can be caused by the genetic factors, including the Mandelian or polygenic inheritance. Traditional methods of identifying genes associated with dyslipidemia are mainly DNA sequencing and linkage analysis, suitable for Mendelian genetic dyslipidemia disease. The rise of next-generation sequencing technology applies to not only Mandelian inheritance, but also complex forms of dyslipidemia diseases. Since 2006, genome-wide association studies (GWAS) screened out many causative genes associated with dyslipidemia, and most of these genes were the previously identified ones by the pedigree-based classic approaches. Furthermore, GWAS revealed that there were the different frequencies of gene variations related to complex forms of dyslipidemia diseases. Most of the identified single nucleotide polymorphisms (SNPs) associated with dyslipidemia are located in non-coding regions and thus people gradually focus on the gene variations of these loci. The identification of the causative genes will provide new insights into the pathophysiology of dyslipidemia diseases and a step toward therapeutic intervention. This review summarized recent pro-gress in heritable dyslipidemia.

Computational evaluation of the axis-blade angle for measurements of implant positions in trochanteric hip fractures: A finite element analysis.

BACKGROUND: Recently, a novel approach axis-blade angle (ABA) was developed to measure implant positions during trochanteric hip fracture surgery. It was defined as the sum of two angles α and ß measured between the femoral neck axis and helical blade axis in anteroposterior and lateral X-ray films, respectively. Although its clinical practicability has been confirmed, the mechanism is yet to be investigated by means of finite element (FE) analysis. METHODS: Computed tomography images of four femurs and dimensions of one implant at three angles were obtained to construct FE models. For each femur, 15 FE models in an arrangement (intramedullary nails at three angles multiplying five blade positions) were established. Under the simulation of normal walking loads, the ABA, von Mises stress (VMS), maximum/minimum principal strain and displacement were analyzed. RESULTS: When the ABA increased, all outcome indicators initially decreased till reaching inferior-middle site and then increased while the blade positions within the femoral head shifted from the superior-anterior quadrant toward the inferior-posterior quadrant, where the ABA were higher. Only the peak VMS of implant models in the inferior-posterior quadrant (particularly the inferior-middle site within) with blades in did not reach the yielding (risky) cut-off. CONCLUSIONS: From the perspective of angles, ABA, this study demonstrated the inferior-posterior quadrant as the relatively stable and safe regions, especially the inferior-middle site within. This was similar but more elaborate compared with previous studies and clinical practice. Therefore, ABA could be employed as a promising approach to anchor the implants into the optimal region.

Reconstruction of Epidural Fat to Prevent Epidural Fibrosis After Laminectomy in Rabbits.

Laminectomy can effectively decompress the spinal cord and expand the vertebral canal. However, the fibrosis that appears may cause adherence and recompression of the spinal cord or/and nerve root, which may cause failed back syndrome (FBS) and make the reexposure process more difficult. Reconstruction of the epidural fat may be an ideal method to achieve satisfactory results. Thirty-six New Zealand rabbits were randomly divided into three groups: control, extracellular matrix (ECM), and ECM+aMSCs groups. Saline, ECM gel, and ECM+aMSC complex were placed, respectively, at the fifth lumbar vertebrate of the rabbits. Epidural fat and fibrosis formation were detected by magnetic resonance imaging (MRI) and histologically at the 4th, 8th, and 12th weeks. Quantitative RT-PCR was used to detect the expression of interleukin 6 (IL-6) and transforming growth factor ß (TGF-ß). MRI and Oil Red O staining revealed epidural fat formation at the 12th week in the ECM+aMSCs group. Hematoxylin and eosin staining showed that the numbers of fibroblasts in the ECM gel and ECM+aMSCs groups were less than the control group at the 4th and 8th weeks (p < 0.05). Masson's trichrome staining showed that the proportion of collagen fibers in the ECM gel and ECM+aMSCs group was lower than the control group (p < 0.05). Quantitative RT-PCR showed the expressions of TGF-ß and IL-6 were lower in the ECM gel and ECM+aMSCs group than those in the control group (p < 0.05) at the 4th week, but higher at the 8th week. We successfully reconstructed the epidural fat with ECM gel and aMSC complex; additionally, IL-6 and TGF-ß cytokines were lower at early stage after laminectomy. Impact statement Our study revealed that epidural fat formed at the 12th week in the extracellular matrix (ECM) plus mesenchymal stem cell (MSC) group after laminectomy in rabbits; additionally, transforming growth factor ß (TGF-ß) (fibrosis) and interleukin 6 (IL-6) (inflammation) expression was reduced. Thus, we believe that our study makes a significant contribution to the literature because we were able to successfully reconstruct the epidural fat with an ECM gel combined with MSCs and reduce local inflammation.

Clinical and molecular cytogenetic findings and pregnancy outcomes of fetuses with isochromosome Y.

BACKGROUND: The mosaic forms and clinical phenotypes of fetuses with isochromosome Y are difficult to predict. Therefore, we summarized the cases of nine fetuses with isochromosome Y identified in prenatal diagnosis with a combination of molecular cytogenetic techniques, providing clinical evidence for prenatal genetic counseling. METHODS: The prenatal diagnosis and pregnancy outcomes of nine fetuses with isochromosome Y were obtained by a  retrospective analysis. Isochromosome Y was identified prenatally by different approaches, such as conventional karyotyping, chromosomal microarray analysis (CMA), quantitative fluorescent polymerase chain reaction (QF-PCR) and fluorescence in situ hybridization (FISH). RESULTS: Seven idic(Y) fetuses and two i(Y) fetuses were identified. One fetus was complete for i(Y)(p10), and the rest with 45,X had mosaic forms. A break and fusion locus was identified in Yp11.3 in one fetus, in Yq11.22 in six fetuses and in Yp10 in two fetuses. The CMA results suggested that different deletions and duplications were found on the Y chromosome. The deletion fragments ranged from 4.7 Mb to the entire Y chromosome, and the duplication fragments ranged from 10.4 to 18.0 Mb. QF-PCR analysis suggested that the AZF region was intact in one fetus, four fetuses had AZFb+c+d deletion, one fetus had AZFa+b+c+d deletion, and one fetus had AZFc+d deletion. Finally, four healthy male neonates were delivered successfully, but the parents of the remaining five fetuses, including three healthy and two unhealthy fetuses, chose to terminate their pregnancies. CONCLUSION: The fetus and neonate phenotype of prenatally detected isochromosome Y usually is that of a normally developed male, ascertained in the absence of other indicators of a fetal structural anomaly. Our study provides clinical reference materials for risk assessment and permits better prenatally counseling and preparation of parents facing the birth of isochromosome Y fetuses.

Exosomal let-7f-5p derived from mineralized osteoblasts promotes the angiogenesis of endothelial cells via the DUSP1/Erk1/2 signaling pathway.

Blood vessel formation is the prerequisite for the survival and growth of tissue-engineered bone. Mineralized osteoblasts (MOBs) have been shown to regulate angiogenesis through the secretion of exosomes containing various pro-angiogenic factors. However, whether the mineralized osteoblast-derived exosomes (MOB-Exos) containing let-7f-5p can regulate the angiogenesis of endothelial cells (ECs) is still unknown. In this study, the angiogenic capabilities of ECs respectively treated with MOB-Exos, let-7f-5p mimicked MOB-Exos (miR mimic group), and let-7f-5p inhibited MOB-Exos (miR inhibitor group) were compared through in vitro and in vivo studies. Moreover, the potential mechanism of MOB-Exo let-7f-5p regulating angiogenesis was explored by verifying the role of the Erk1/2 signaling pathway and target gene DUSP1. The results showed that MOB-Exos could significantly promote the angiogenesis of ECs, which could be enhanced by mimicked exosomal let-7f-5p and attenuated by inhibited exosomal let-7f-5p. Let-7f-5p could suppress the luciferase activity of wide-type DUSP1, and the mutation of DUSP1 could abrogate the repressive ability of let-7f-5p. Furthermore, the expression of DUSP1 exhibited a reversed trend to that of pErk1/2. The expression of pErk1/2 was significantly higher in the miR mimic group and lower in the miR inhibitor group than that in the MOB-Exos group, while inhibition of pErk1/2 could partly impair the angiogenic capabilities of ECs. In conclusion, we concluded that exosomal let-7f-5p derived from MOBs could promote the angiogenesis of ECs via activating the DUSP1/Erk1/2 signaling pathway, which might be a promising target for promoting the angiogenesis of tissue-engineered bone.

When to Reduce and Fix Displaced Lesser Trochanter in Treatment of Trochanteric Fracture: A Systematic Review.

Purpose: To systematically evaluate the benefits of reducing and fixing displaced lesser trochanter (LT) of trochanteric fractures and when this procedure is worth the effect. Methods: From database establishment through March 2021, four online databases (PubMed, Cochrane, Embase, and Web of Science) were searched for relevant literature that investigated reduction and fixation for displaced LT of trochanteric fractures. The papers were then screened by two reviewers independently and in duplicate according to prior inclusion and exclusion criteria. Demographic data as well as data on fracture types, surgical protocols, and surgical outcomes were recorded, analyzed, and interpreted. Results: Total 10 clinical studies with 928 patients were included, in which 48 cases had intact LT and 880 cases involved the displaced LT, of which 196 (22.27%) cases underwent reduction and fixation for LT while the rest of 684 (77.73%) cases not. In these studies, complications were evaluated as a more applicable predictive parameter for operation than postoperative hip function. Conclusion: It was beneficial to reduce and fix the displaced LT when one of the conditions below occurred: displacement distance of LT ≥2 cm, quantity of comminuted LT fragments ≥2, and range of LT fragments in medial wall ≥75%; the fracture line of LT fragments reaching or exceeding the midline of the posterior wall.

No significant association between RELN polymorphism and autism in case-control and family-based association study in Chinese Han population.

The present study genotyped four SNPs (rs736707, rs2229864, rs362691, and rs2073559) of the Reelin gene (RELN) in 165 autistic trios, 67 sporadic autistic children and 283 healthy controls with Chinese Han pedigree. Both case-control analysis and transmission disequilibrium test (TDT) found no evidence of significant association. The results do not support previous positive findings and suggest that the four single-nucleotide polymorphisms (SNP) of RELN are unlikely to be associated with childhood autism in Chinese Han population.

Cyclic pulsation stress promotes bone formation of tissue engineered laminae through the F-actin/YAP-1/ß-Catenin signaling axis.

Mechanical loads are fundamental regulators of bone formation and remodeling. However, the molecular regulation of mechanotransduction during vertebral laminae regeneration remains poorly understood. Here, we found that cerebrospinal fluid pulsation (CSFP) stress-cyclic pulsation stress-could promote the osteogenic and angiogenic abilities of rat mesenchymal stromal cells (MSC), thereby promoting tissue-engineered laminae's bone and blood vessel formation. In the process, F-actin relayed CSFP stress to promote the nuclear translocation of YAP1, which then decreased the degradation and promoted the nuclear translocation of ß-Catenin. In turn, the nuclear translocation of ß-Catenin promoted the osteogenic differentiation and angiogenic abilities of MSC, thereby promoting tissue-engineered laminae's bone and blood vessel formation. Thus, we conclude that CSFP promotes the osteogenesis and angiogenesis of tissue-engineered laminae through the F-actin/YAP-1/ß-Catenin signaling axis. This study advances our understanding of vertebral laminae regeneration and provides potential therapeutic approaches for spinal degeneration after spinal laminectomy.

Wnt/ß-Catenin Pathway Balances Scaffold Degradation and Bone Formation in Tissue-Engineered Laminae.

Tissue engineering provides a promising way for the regeneration of artificial vertebral laminae. Previous studies have confirmed the feasibility of reconstructing vertebral laminae via hydroxyapatite-collagen I scaffolds and mesenchymal stromal cells. However, there were no studies exploring the degradation of hydroxyapatite-collagen I scaffolds and the function of Wnt/ß-catenin pathway in the process. In this study, tissue-engineered laminae (TEL) were constructed by nanohydroxyapatite/collagen I scaffolds and umbilical cord Wharton's Jelly mesenchymal stromal cells (WJ-MSCs). Cell attachment was observed by scanning electron microscopy, and cell viability was confirmed by Live/Dead staining. The rat models were randomly divided into control and ß-catenin inhibition groups. Vertebral lamina defect rat models were made on the fifth lumbar vertebrate, and TEL was implanted into the defect site. After 14 weeks, the newborn laminae were harvested for microcomputed tomography, histology, or transcriptional profile analysis. We found that, for the control group, the newborn lamina formation matched with the scaffold degradation and complete newborn laminae formed at the 14th week; for the ß-catenin inhibition group, the scaffold degradation rate overrated the lamina formation and no complete artificial laminae were formed at the 14th week. In addition, the osteoclastic genes, such as Cathepsin K or RANKL, in the control groups were significantly lower than the ß-catenin inhibition group, and the antiosteoclastic gene, OPG, in the control group was significantly higher than the ß-catenin inhibition group. In conclusion, inhibition of Wnt/ß-catenin pathway led to speedy scaffold degradation and deferred artificial lamina formation. Wnt/ß-catenin pathway played a critical role in maintaining the balance between scaffold degradation and bone formation in the process of vertebral lamina reconstruction.

Cerebrospinal Fluid Pulsation Stress Promotes the Angiogenesis of Tissue-Engineered Laminae.

BACKGROUND: Angiogenesis is a prerequisite step to achieve the success of bone regeneration by tissue engineering technology. Previous studies have shown the role of cerebrospinal fluid pulsation (CSFP) stress in the reconstruction of tissue-engineered laminae. In this study, we investigated the role of CSFP stress in the angiogenesis of tissue-engineered laminae. METHODS: For the in vitro study, a CSFP bioreactor was used to investigate the impact of CSFP stress on the osteogenic mesenchymal stem cells (MSCs). For the in vivo study, forty-eight New Zealand rabbits were randomly divided into the CSFP group and the Non-CSFP group. Tissue-engineered laminae (TEL) was made by hydroxyapatite-collagen I scaffold and osteogenic MSCs and then implanted into the lamina defect in the two groups. The angiogenic and osteogenic abilities of newborn laminae were examined with histological staining, qRT-PCR, and radiological analysis. RESULTS: The in vitro study showed that CSFP stress could promote the vascular endothelial growth factor A (VEGF-A) expression levels of osteogenic MSCs. In the animal study, the expression levels of angiogenic markers in the CSFP group were higher than those in the Non-CSFP group; moreover, in the CSFP group, their expression levels on the dura mater surface, which are closer to the CSFP stress stimulation, were also higher than those on the paraspinal muscle surface. The expression levels of osteogenic markers in the CSFP group were also higher than those in the Non-CSFP group. CONCLUSION: CSFP stress could promote the angiogenic ability of osteogenic MSCs and thus promote the angiogenesis of tissue-engineered laminae. The pretreatment of osteogenic MSC with a CSFP bioreactor may have important implications for vertebral lamina reconstruction with a tissue engineering technique.