Delta Cortisol and Adrenal Insufficiency

The ACTH stimulation test is used to diagnose adrenal insufficiency (AI). This study evaluated the diagnostic performance of serum delta cortisol from ACTH stimulation tests and determined appropriate cut-off levels of serum 30-minute delta cortisol for the diagnosis of AI, allowing a reduction in the number of 60-minute cortisol tests. A 6-year retrospective study in 471 patients was conducted. The performance of the serum delta cortisol in diagnosing AI was assessd using a multivariable logistic regression model and the area under ROC curves (AuROC). Both serum 30-minute and 60-minute delta cortisol demonstrated equally high diagnostic accuracy for AI (AuROC for LDT : 0.91 vs 0.90 ; HDT : 0.91 vs 0.92, respectively). The 30-minute delta cortisol test was chosen to develop proposed diagnostic cut-off levels due to its simplicity. The proposed lower cut-off level for 30-minute delta cortisol was Δ 11.8 μg/dL for LDT and Δ > 10.5 μg/dL for HDT. These cut-off levels yielded high sensitivity and specificity > 90%. The 30-minute serum delta cortisol using the proposed cut-off levels provides diagnostic performance for AI equal to that of the 60-minute test and is more convenient, requires less time, less invasive and is cost-saving

Identifying Chemical Composition, Safety and Bioactivity of Thai Rice Grass Extract Drink in Cells and Animals

From MDPI via Jisc Publications RouterHistory: accepted 2021-11-11, pub-electronic 2021-11-15Publication status: PublishedFunder: the Vejadusit Foundation under the Royal Patronage of His Royal Highness Princess Kanlyani Wattana, Bangkok Hospital, Thailand.; Grant(s): 08/07/2558Rice grass has been reported to contain bioactive compounds that possess antioxidant and free-radical scavenging activities. We aimed to assess rice grass extract (RGE) drink by determining catechin content, free-radical scavenging and iron-binding properties, as well as toxicity in cells and animals. Young rice grass (Sukhothai-1 strain) was dried, extracted with hot water and lyophilized in a vacuum chamber. The resulting extract was reconstituted with deionized water (260 mg/40 mL) and served as Sukhothai-1 rice grass extract drink (ST1-RGE). HPLC results revealed at least eight phenolic compounds, for which the major catechins were catechin, epicatechin and epigallocatechin-3-gallate (EGCG) (2.71–3.57, 0.98–1.85 and 25.47–27.55 mg/40 mL serving, respectively). Elements (As, Cu, Pb, Sn and Zn) and aflatoxin (B1, B2, G1 and G2) contents did not exceed the relevant limits when compared with WHO guideline values. Importantly, ST1-RGE drink exerted radical-scavenging, iron-chelating and anti-lipid peroxidation properties in aqueous and biological environments in a concentration-dependent manner. The drink was not toxic to cells and animals. Thus, Sukhothai-1 rice grass product is an edible drink that is rich in catechins, particularly EGCG, and exhibited antioxidant, free radical scavenging and iron-binding/chelating properties. The product represents a functional drink that is capable of alleviating conditions of oxidative stress and iron overload

Differentiation-dependent association of phosphorylated extracellular signal-regulated kinase with the chromatin of osteoblast-related genes

The ERK/MAP kinase pathway is an important regulator of gene expression and differentiation in postmitotic cells. To understand how this pathway controls gene expression in bone, we examined the subnuclear localization of P-ERK in differentiating osteoblasts. Induction of differentiation was accompanied by increased ERK phosphorylation and expression of osteoblast-related genes, including osteocalcin ( Bglap2 ) and bone sialoprotein ( Ibsp ). Confocal immunofluorescence microscopy revealed that P-ERK colocalized with the RUNX2 transcription factor in the nuclei of differentiating cells. Interestingly, a portion of this nuclear P-ERK was directly bound to the proximal promoter regions of Bglap2 and Ibsp . Furthermore, the level of P-ERK binding to chromatin increased with differentiation, whereas RUNX2 binding remained relatively constant. The P-ERK-chromatin interaction was seen only in RUNX2-positive cells, required intact RUNX2-selective enhancer sequences, and was blocked with MAPK inhibition. These studies show for the first time that RUNX2 specifically targets P-ERK to the chromatin of osteoblast-related genes, where it may phosphorylate multiple substrates, including RUNX2, resulting in altered chromatin structure and gene expression. © 2010 American Society for Bone and Mineral ResearchPeer Reviewedhttp://deepblue.lib.umich.edu/bitstream/2027.42/64899/1/90705_ftp.pd

miRNA-Mediated Functional Changes through Co-Regulating Function Related Genes

BACKGROUND: MicroRNAs play important roles in various biological processes involving fairly complex mechanism. Analysis of genome-wide miRNA microarray demonstrate that a single miRNA can regulate hundreds of genes, but the regulative extent on most individual genes is surprisingly mild so that it is difficult to understand how a miRNA provokes detectable functional changes with such mild regulation. RESULTS: To explore the internal mechanism of miRNA-mediated regulation, we re-analyzed the data collected from genome-wide miRNA microarray with bioinformatics assay, and found that the transfection of miR-181b and miR-34a in Hela and HCT-116 tumor cells regulated large numbers of genes, among which, the genes related to cell growth and cell death demonstrated high Enrichment scores, suggesting that these miRNAs may be important in cell growth and cell death. MiR-181b induced changes in protein expression of most genes that were seemingly related to enhancing cell growth and decreasing cell death, while miR-34a mediated contrary changes of gene expression. Cell growth assays further confirmed this finding. In further study on miR-20b-mediated osteogenesis in hMSCs, miR-20b was found to enhance osteogenesis by activating BMPs/Runx2 signaling pathway in several stages by co-repressing of PPARγ, Bambi and Crim1. CONCLUSIONS: With its multi-target characteristics, miR-181b, miR-34a and miR-20b provoked detectable functional changes by co-regulating functionally-related gene groups or several genes in the same signaling pathway, and thus mild regulation from individual miRNA targeting genes could have contributed to an additive effect. This might also be one of the modes of miRNA-mediated gene regulation

Human Umbilical Cord Blood-Derived CD34+ Cells Reverse Osteoporosis in NOD/SCID Mice by Altering Osteoblastic and Osteoclastic Activities

Osteoporosis is a bone disorder associated with loss of bone mineral density and micro architecture. A balance of osteoblasts and osteoclasts activities maintains bone homeostasis. Increased bone loss due to increased osteoclast and decreased osteoblast activities is considered as an underlying cause of osteoporosis.The cures for osteoporosis are limited, consequently the potential of CD34+ cell therapies is currently being considered. We developed a nanofiber-based expansion technology to obtain adequate numbers of CD34(+) cells isolated from human umbilical cord blood, for therapeutic applications. Herein, we show that CD34(+) cells could be differentiated into osteoblastic lineage, in vitro. Systemically delivered CD34(+) cells home to the bone marrow and significantly improve bone deposition, bone mineral density and bone micro-architecture in osteoporotic mice. The elevated levels of osteocalcin, IL-10, GM-CSF, and decreased levels of MCP-1 in serum parallel the improvements in bone micro-architecture. Furthermore, CD34(+) cells improved osteoblast activity and concurrently impaired osteoclast differentiation, maturation and functionality.These findings demonstrate a novel approach utilizing nanofiber-expanded CD34(+) cells as a therapeutic application for the treatment of osteoporosis

Transcriptional Analysis of Fracture Healing and the Induction of Embryonic Stem Cell–Related Genes

Fractures are among the most common human traumas. Fracture healing represents a unique temporarily definable post-natal process in which to study the complex interactions of multiple molecular events that regulate endochondral skeletal tissue formation. Because of the regenerative nature of fracture healing, it is hypothesized that large numbers of post-natal stem cells are recruited and contribute to formation of the multiple cell lineages that contribute to this process. Bayesian modeling was used to generate the temporal profiles of the transcriptome during fracture healing. The temporal relationships between ontologies that are associated with various biologic, metabolic, and regulatory pathways were identified and related to developmental processes associated with skeletogenesis, vasculogenesis, and neurogenesis. The complement of all the expressed BMPs, Wnts, FGFs, and their receptors were related to the subsets of transcription factors that were concurrently expressed during fracture healing. We further defined during fracture healing the temporal patterns of expression for 174 of the 193 genes known to be associated with human genetic skeletal disorders. In order to identify the common regulatory features that might be present in stem cells that are recruited during fracture healing to other types of stem cells, we queried the transcriptome of fracture healing against that seen in embryonic stem cells (ESCs) and mesenchymal stem cells (MSCs). Approximately 300 known genes that are preferentially expressed in ESCs and ∼350 of the known genes that are preferentially expressed in MSCs showed induction during fracture healing. Nanog, one of the central epigenetic regulators associated with ESC stem cell maintenance, was shown to be associated in multiple forms or bone repair as well as MSC differentiation. In summary, these data present the first temporal analysis of the transcriptome of an endochondral bone formation process that takes place during fracture healing. They show that neurogenesis as well as vasculogenesis are predominant components of skeletal tissue formation and suggest common pathways are shared between post-natal stem cells and those seen in ESCs

Osteoimmunology of Oral and Maxillofacial Diseases : Translational Applications Based on Biological Mechanisms

The maxillofacial skeleton is highly dynamic and requires a constant equilibrium between the bone resorption and bone formation. The field of osteoimmunology explores the interactions between bone metabolism and the immune response, providing a context to study the complex cellular and molecular networks involved in oro-maxillofacial osteolytic diseases. In this review, we present a framework for understanding the potential mechanisms underlying the immuno-pathobiology in etiologically-diverse diseases that affect the oral and maxillofacial region and share bone destruction as their common clinical outcome. These otherwise different pathologies share similar inflammatory pathways mediated by central cellular players, such as macrophages, T and B cells, that promote the differentiation and activation of osteoclasts, ineffective or insufficient bone apposition by osteoblasts, and the continuous production of osteoclastogenic signals by immune and local stromal cells. We also present the potential translational applications of this knowledge based on the biological mechanisms involved in the inflammation-induced bone destruction. Such applications can be the development of immune-based therapies that promote bone healing/regeneration, the identification of host-derived inflammatory/collagenolytic biomarkers as diagnostics tools, the assessment of links between oral and systemic diseases; and the characterization of genetic polymorphisms in immune or bone-related genes that will help diagnosis of susceptible individuals.Peer reviewe

Cooperative interactions between bone morphogenetic proteins and the RUNX2 transcription factor in osteoblast differentiation.

Bone morphogenetic proteins (BMPs) are potent osteogenic factors that stimulate osteoblast differentiation and bone formation. BMPs are known to induce expression of RUNX2/CBFA1 and DLX5 transcription factors. Because overexpression of RUNX2 induces osteoblast differentiation and mineralization, mimicking the effects of BMPs, it has been proposed that this factor mediates actions of BMPs in stimulating osteoblast differentiation. However, calvarial-derived cells from Cbfa1-deficient mice still show a modest response to BMPs, suggesting that certain effects of BMPs do not require RUNX2. Two approaches were used to explore whether BMPs and RUNX2 function in a complementary manner: (1) BMP blocking antibodies were used to examine the effect of BMPs on RUNX2-mediated osteoblast differentiation; (2) combined treatment with recombinant human BMP2 (rhBMP2) and RUNX2 was used to examine complementary cellular responsiveness to these two factors. Our results clearly indicate that BMPs are required for RUNX2-dependent transcription and induction of osteoblast differentiation markers, and in turn RUNX2 enhances cellular responsiveness to BMPs as documented by the synergistic induction of osteoblast differentiation by BMP2 and RUNX2. All these pieces of evidence establish the complementary function between BMPs and RUNX2. We further examined the mechanism responsible for this phenomenon. Interestingly, even though RUNX2 enhances cellular responsiveness to BMP2, it did not affect the ability of BMP2 to stimulate downstream signal transduction pathways (SMAD1/5/8, p38, JNK and ERK). In contrast, BMP2 and RUNX2 synergistically induced Dlx5 expression and this induction preceded induction of Ocn and Bsp genes. More interestingly, DLX5 cooperatively functions with SMAD1/4 in stimulating Ocn and Bsp , and this cooperative function occurs via protein-protein interactions. In conclusion, our results indicate that the mechanism responsible for the interplay between BMPs and RUNX2 occurs after receptor activation. Furthermore, they suggest that DLX5 functions as a primary gene for stimulating Ocn and Bsp genes, and the cooperative function between DLX5 and SMAD1/4 may lead to the complementary function between BMPs and RUNX2.Ph.D.BiochemistryBiological SciencesCellular biologyMolecular biologyPure SciencesUniversity of Michigan, Horace H. Rackham School of Graduate Studieshttp://deepblue.lib.umich.edu/bitstream/2027.42/125736/2/3208533.pd

Identifying Chemical Composition, Safety and Bioactivity of Thai Rice Grass Extract Drink in Cells and Animals

Rice grass has been reported to contain bioactive compounds that possess antioxidant and free-radical scavenging activities. We aimed to assess rice grass extract (RGE) drink by determining catechin content, free-radical scavenging and iron-binding properties, as well as toxicity in cells and animals. Young rice grass (Sukhothai-1 strain) was dried, extracted with hot water and lyophilized in a vacuum chamber. The resulting extract was reconstituted with deionized water (260 mg/40 mL) and served as Sukhothai-1 rice grass extract drink (ST1-RGE). HPLC results revealed at least eight phenolic compounds, for which the major catechins were catechin, epicatechin and epigallocatechin-3-gallate (EGCG) (2.71–3.57, 0.98–1.85 and 25.47–27.55 mg/40 mL serving, respectively). Elements (As, Cu, Pb, Sn and Zn) and aflatoxin (B1, B2, G1 and G2) contents did not exceed the relevant limits when compared with WHO guideline values. Importantly, ST1-RGE drink exerted radical-scavenging, iron-chelating and anti-lipid peroxidation properties in aqueous and biological environments in a concentration-dependent manner. The drink was not toxic to cells and animals. Thus, Sukhothai-1 rice grass product is an edible drink that is rich in catechins, particularly EGCG, and exhibited antioxidant, free radical scavenging and iron-binding/chelating properties. The product represents a functional drink that is capable of alleviating conditions of oxidative stress and iron overload