BACKGROUND: Colorectal cancer (CRC) remains a major global health challenge, with high incidence and mortality rates. The role of long noncoding RNAs (lncRNAs) in cancer progression has received considerable attention. The present study aimed to investigate the function and mechanisms underlying the role of lncRNA RP11-197K6.1, microRNA-135a-5p (hsa-miR-135a-5p), and DLX5 in CRC development.
METHODS: We analyzed RNA sequencing data from The Cancer Genome Atlas Colorectal Cancer dataset to identify the association between lncRNA RP11-197K6.1 and CRC progression. The expression levels of lncRNA RP11-197K6.1 and DLX5 in CRC samples and cell lines were determined by real-time quantitative PCR and western blotting assays. Fluorescence in situ hybridization was used to confirm the cellular localization of lncRNA RP11-197K6.1. Cell migration capabilities were assessed by Transwell and wound healing assays, and flow cytometry was performed to analyze apoptosis. The interaction between lncRNA RP11-197K6.1 and miR-135a-5p and its effect on DLX5 expression were investigated by the dual-luciferase reporter assay. Additionally, a xenograft mouse model was used to study the in vivo effects of lncRNA RP11-197K6.1 on tumor growth, and an immunohistochemical assay was performed to assess DLX5 expression in tumor tissues.
RESULTS: lncRNA RP11-197K6.1 was significantly upregulated in CRC tissues and cell lines as compared to that in normal tissues, and its expression was inversely correlated with patient survival. It promoted the migration and metastasis of CRC cells by interacting with miR-135a-5p, alleviated suppression of DLX5 expression, and facilitated tumor growth.
CONCLUSIONS: This study demonstrated the regulatory network and mechanism of action of the lncRNA RP11-197K6.1/miR-135a-5p/DLX5 axis in CRC development. These findings provided insights into the molecular pathology of CRC and suggested potential therapeutic targets for more effective treatment of patients with CRC.

Retroviral-mediated misexpression in chicken embryos has been a powerful research tool for developmental biologists in the last two decades. In the RCASBP retroviral vectors that are widely used for in vivo somatic transgenesis, a coding sequence of interest is under the transcriptional control of a strong viral promoter in the long terminal repeat. While this has proven to be effective for studying secreted signalling proteins, interpretation of the mechanisms of action of nuclear factors is more difficult using this system since it is not clear whether phenotypic effects are cell-autonomous or not, and therefore whether they represent a function of the endogenous protein. Here, we report the consequences of retroviral expression using the RCANBP backbone, in which the transcription factor Dlx5 is expressed under the control of chondrocyte-specific regulatory sequences from the Col2a1 gene. To our knowledge, this is the first demonstration of a tissue-specific phenotype in the chicken embryo.

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BACKGROUND: Preeclampsia is a hypertensive disorder of pregnancy. DLX5 plays an important role in the migration and differentiation of subglobus pallidus precursor cells.
METHODS: We established a zebrafish line expressing high levels of DLX5 and investigated changes in behavior and development of the nervous system.
RESULTS: The ratios of brain volume area to whole body area at 96 hpf zebrafish in the experimental group (gRNA + CasRx) were significantly lower than the WT group and the negative control group (casRx) (P < 0.01). Behavioral trajectory distances and movement speeds exhibited by the 6th day of development in zebrafish in the experimental group (gRNA + CasRx) were significantly shorter (P < 0.01) and lower (P < 0.05) than the negative control group (gRNA + CasRx), respectively.
CONCLUSIONS: Data suggested that the increased expression levels of DLX5 can inhibit brain volume development and behavioral activities in zebrafish. Maybe the high expression levels of DLX5 in the pathological state of preeclampsia can inhibit the development of the nervous system in offspring.

Despite significant progress in our understanding of the molecular mechanism of mesenchymal stem cell (MSC) differentiation, less is known about the factors maintaining the stemness and plasticity of MSCs. Here, we show that the NFIB-MLL1 complex plays key roles in osteogenic differentiation and stemness of C3H10T1/2 MSCs. We find that depletion of either NFIB or MLL1 results in a severely hampered osteogenic potential and failed activation of key osteogenic transcription factors, such as Dlx5, Runx2, and Osx, following osteogenic stimuli. In addition, the NFIB-MLL1 complex binds directly to the promoter of Dlx5, and exogenous expression of Myc-Dlx5, but not the activation of either the BMP- or the Wnt-signaling pathway, is sufficient to restore the osteogenic potential of cells depleted of NFIB or MLL1. Moreover, chromatin immunoprecipitation (ChIP) and ChIP-sequencing analysis showed that the NFIB-MLL1 complex mediates the deposition of trimethylated histone H3K4 at both Dlx5 and Cebpa, key regulator genes that function at the early stages of osteogenic and adipogenic differentiation, respectively, in uncommitted C3H10T1/2 MSCs. Surprisingly, the depletion of either NFIB or MLL1 leads to decreased trimethylated histone H3K4 and results in elevated trimethylated histone H3K9 at those developmental genes. Furthermore, gene expression profiling and ChIP-sequencing analysis revealed lineage-specific changes in chromatin landscape and gene expression in response to osteogenic stimuli. Taken together, these data provide evidence for the hitherto unknown role of the NFIB-MLL1 complex in the maintenance and lineage-specific differentiation of C3H10T1/2 MSCs and support the epigenetic regulatory mechanism underlying the stemness and plasticity of MSCs.

Split Hand-Foot Malformation (SHFM) is a congenital limb defect characterized by a median cleft of the hands and/or feet due to the absence/hypoplasia of the central rays. It may occur as part of a syndromic condition or as an isolated malformation. The most common of the six genetic loci identified for this condition is correlated to SHFM1 and maps in the 7q21q22 region. SHFM1 is characterized by autosomal dominant transmission, incomplete penetrance and variable expressivity. Associated features often include hearing loss, intellectual disability/developmental delay and craniofacial abnormalities. Disruption of the DLX5/DLX6 genes, mapping within the SHFM1 locus, is now known to be responsible for the phenotype. Through SNP array, we analyzed a patient affected by SHFM1 associated with deafness and an abnormality of the inner ear (incomplete partition type I); we identified a deletion in 7q21, not involving the DLX5/6 genes, but including exons 15 and 17 of DYNC1I1, known to act as exonic enhancers (eExons) of the DLX5/6 genes. We further demonstrated the role of DYNC1I1 eExons in regulating DLX5/6 expression by means of showing a reduced expression of the DLX5/6 genes through RT-PCR in a patient-derived lymphoblastoid cell line. Furthermore, our data and a review of published cases do not support the hypothesis that DLX5/6 are imprinted in humans. This work is an example of how the disruption of regulatory elements can be responsible for congenital malformations.

High-throughput RNA-sequencing can determine the impact of nutrients and their combinations on gene transcription levels in osteocytes, and clarify the biological pathways associated with their impact on bone tissues. Previously, we reported that resveratrol (RES) and peonidin-3-O-glucoside (POG) increased osteoblastogenesis, as well as reduced osteoclastogenesis in transgenic teleost fish models. Here, we perform whole-genome transcriptomic profiling of osteoblasts treated with POG or RES to provide a comprehensive understanding of alterations in gene expression and the molecular mechanisms involved. Cultured human fetal osteoblastic hFOB 1.19 cells were treated with the test compounds, and then RNA was used to prepare RNA-seq libraries, that were sequenced using a NovaSeq 6000. Treatment with POG or RES increased osteoblast proliferation and reduced apoptosis. Transcriptomic profiling showed that of the 29,762 genes investigated, 3177 were differentially expressed (1481 upregulated, 1696 downregulated, FDR ≤ 0.05) in POG-treated osteoblasts. In the RES-treated osteoblasts, 2288 genes were differentially expressed (DGEs, 1068 upregulated, 1220 downregulated, FDR ≤ 0.05). Ingenuity® Pathway Analysis (IPA) of DGEs from RES or POG-treated osteoblasts revealed significant downregulation of the apoptosis, osteoarthritis and HIF1α canonical pathways, and a significant reduction in Rankl mRNA expression. The data suggest that RES and POG have both anabolic and anticlastogenic effects.

Osteoarthritis (OA) has been considered non-reversible as articular cartilage wears down with limited repair capacity. Enhanced chondrocyte hypertrophy and increased type X collagen gene (COL10A1) expression have been associated with OA. Therefore, regulators controlling collagen X expression and chondrocyte hypertrophy may play a role in OA intervention. Here, we investigated how Distal-less homeobox 5 (DLX5), the distal-less homeobox family member, controls murine Col10a1 gene expression and chondrocyte hypertrophy in chondrogenic cell models and its role in a murine OA model. Through qRT-PCR and Western blot analyses, we detected significantly increased levels of COL10A1 and DLX5 in hypertrophic MCT and ATDC5 cells compared to their proliferative stage. Forced expression of Dlx5 further increases, while knockdown of Dlx5 decreases COL10A1 expression in hypertrophic MCT cells. We have performed dual-luciferase reporter and ChIP assays and demonstrated that DLX5 promotes reporter activity through direct interaction with Col10a1 cis-enhancer. We established a murine OA model and detected markedly increased COL10A1 and DLX5 in the articular cartilage and subchondral bone of the OA mice compared with the controls. Notably, forced overexpression of DLX5 in hypertrophic MCT cells up-regulates RUNX2, and adjacent DLX5 and RUNX2 binding sites have previously been found within the Col10a1 cis-enhancer. Together, our data suggest that DLX5 may cooperate with RUNX2 to control cell-specific Col10a1 expression and chondrocyte hypertrophy and is involved in OA pathogenesis.

UNASSIGNED: Ossification of the posterior longitudinal ligaments (OPLL) is common disorder characterized by heterotopic ossification of the spinal ligaments. Mechanical stimulation (MS) plays an important role in OPLL. DLX5 is an essential transcription factor required for osteoblast differentiation. However, the role of DLX5 during in OPLL is unclear. This study aims to investigate whether DLX5 is associated with OPLL progression under MS.
UNASSIGNED: Stretch stimulation was applied to spinal ligaments cells derived from OPLL (OPLL cells) and non-OPLL (non-OPLL cells) patients. Expression of DLX5 and osteogenesis-related genes were determined by quantitative real-time polymerase chain reaction and Western blot. The osteogenic differentiation ability of the cells was measured using alkaline phosphatase (ALP) staining and alizarin red staining. The protein expression of DLX5 in the tissues and the nuclear translocation of NOTCH intracellular domain (NICD) was examined by immunofluorescence.
UNASSIGNED: Compared with non-OPLL cells, OPLL cells expressed higher levels of DLX5 in vitro and vivo (p < 0.01). Upregulated expression of DLX5 and osteogenesis-related genes (OSX, RUNX2, and OCN) were observed in OPLL cells induced with stretch stimulation and osteogenic medium, whereas there was no change in the non-OPLL cells (p < 0.01). Cytoplasmic NICD protein translocated from the cytoplasm to the nucleus inducing DLX5 under stretch stimulation, which was reduced by the NOTCH signaling inhibitors (DAPT) (p < 0.01).
UNASSIGNED: These data suggest that DLX5 play a critical role in MS-induced progression of OPLL through NOTCH signaling, which provides a new insight into the pathogenesis of OPLL.

UNASSIGNED: Authors report a rare case report about split hand and foot malformation (SHFM) also sometimes referred to as ectrodactyly.
UNASSIGNED: The patient with hand and foot malformations presented to casualty. A 60-year-old male was brought with alleged history of road traffic accident with tenderness and deformity in left thigh. On further physical examination, a malformation was present in bilateral feet and right hand. Plain radiographs were taken after emergency primary management which revealed a fracture of shaft of femur of the left side and absence of 2nd and 3rd phalanges in bilateral feet and lobster claw like malformation in the right hand. The patient was further investigated and operated with femur interlocking nail and later discharged under stable condition. Screening for other congenital defects was done.
UNASSIGNED: Patients with SHFM should undergo screening for other congenital anomalies. Electrocardiogram, 2D ECHO, chest radiograph, and ultrasonography abdomen should be done. Genetic analysis ideally should be done to identify mutations involved. Surgical intervention is only required when patient demands improved function of limb.