Expression Of Parathyroid Hormone-like Hormone Research Articles

Streptonigrin Mitigates Lung Cancer-induced Cachexia by Suppressing TCF4/TWIST1-induced PTHLH Expression.

Cachexia - a wasting disorder of adipose and skeletal muscle tissue - is the most common driver of poor prognosis in patients with advanced lung cancer. Parathyroid hormone-like hormone (PTHLH) is potentially a critical factor in cancer-associated cachexia. We previously showed that streptonigrin - an aminoquinone with antitumor effects - inhibited the interaction between TCF4 and TWIST1. This study aimed to determine the anti-cachectic performance of streptonigrin in lung cancer. We assessed the effect of streptonigrin on the interaction of TCF4 and TWIST1 using co-immunoprecipitation and a mammalian-two hybrid luciferase assay, which was confirmed by an in vitro GST pull-down assay using recombinant bHLH domain-containing TCF4 and TWIST1. We assessed the anti-cachectic effect of streptonigrin in vivo using an LLC1 cell-induced tumour-bearing mouse model. Changes in the degree of skeletal muscle and adipose tissue wasting were determined by measuring the weights of gastrocnemius and epidydimal white adipose tissue. Streptonigrin was found to inhibit the interaction of TCF4 with TWIST1 in a dose-dependent manner. The in vitro GST pull-down assay revealed that streptonigrin directly inhibited the interaction between TCF4 and TWIST1. The expression of PTHLH mRNA, which is transcriptionally regulated by the TCF4/TWIST1 complex in response to TGF-β1 signalling, was decreased in streptonigrin-treated lung cancer cells. Streptonigrin significantly decreased the expression of proteolysis-related genes in skeletal muscle and browning-related genes in white adipose tissues of LLC1-induced tumour-bearing mice. Streptonigrin exerts potent therapeutic effects on lung cancer-induced cachexia by suppressing TCF4/TWIST1-mediated PTHLH expression.

CEP55 3'-UTR promotes epithelial-mesenchymal transition and enhances tumorigenicity of bladder cancer cells by acting as a ceRNA regulating miR-497-5p.

Centrosomal protein 55 (CEP55) is implicated in the tumorigenesis of bladder cancer (BC) but the detailed molecular mechanisms are unknown. We aim to develop a potential competing endogenous RNA (ceRNA) network related with CEP55 in BC. We first extracted the expression profiles of RNAs from The Cancer Genome Atlas (TCGA) database and used bioinformatic analysis to establish ceRNAs in BC. Real-time quantity PCR (RT-qPCR) and immunohistochemical analysis were performed to measure CEP55 expression in different bladder cell lines and different grades of cancer. Bioinformatics analysis and luciferase assays were conducted to predict potential binding sites among miR-497-5p, CEP55, parathyroid hormone like hormone (PTHLH) and high mobility group A2 (HMGA2). Tumor xenograft model was used to show the effect of CEP55 3'-UTR on cisplatin therapy. Bioinformatics analysis, luciferase assays, and 5' rapid amplification of cDNA ends (5'RACE) were to explore the function of CEP55 3'-untranslated region (3'-UTR) on targeting miR-497-5p. Western blot and immunofluorescence assays were to detect the epithelial-mesenchymal transition (EMT) induction of CEP55 3'-UTR. CEP55 expression as well as the expression levels of the oncogenic proteins PTHLH and HMGA2 were upregulated in BC cells while miR-497-5p was downregulated. Low miR-497-5p expression and high CEP55 and HMGA2 expression levels were associated with more advanced tumor clinical stage and pathological grade. Overexpression of the CEP55 3'-UTR promoted the proliferation, migration, and invasion of the EJ cell line in vitro and accelerated EJ-derived tumor growth in nude mice, while inhibition of the CEP55 3'-UTR suppressed all of these oncogenic processes. In addition, CEP55 3'-UTR upregulation reduced the cisplatin sensitivity of BC cell lines and xenograft tumors. Bioinformatics analysis, luciferase assays, and 5'RACE suggested that the CEP55 3'-UTR functions as a ceRNA targeting miR-497-5p, leading to miR-497-5p downregulation and disinhibition of PTHLH and HMGA2 expression. Further, CEP55 downregulated miR-497-5p transcription by promoting NF-[Formula: see text]B signaling. In turn, CEP55 3'-UTR ultimately promotes EMT and tumorigenesis by activating P38MAPK and ERK 1/2 pathways. These results suggest that a ceRNA regulatory network involving CEP55 upregulates PTHLH and HMGA2 expression by suppressing endogenous miR-497-5p. We unveiled a novel mechanism of BC metastasis, and could become novel therapeutics targets in BC.

Polygonum cuspidatum Extract (Pc-Ex) Containing Emodin Suppresses Lung Cancer-Induced Cachexia by Suppressing TCF4/TWIST1 Complex-Induced PTHrP Expression.

Cachexia, which is characterised by the wasting of fat and skeletal muscles, is the most common risk factor for increased mortality rates among patients with advanced lung cancer. PTHLH (parathyroid hormone-like hormone) is reported to be involved in the pathogenesis of cancer cachexia. However, the molecular mechanisms underlying the regulation of PTHLH expression and the inhibitors of PTHLH have not yet been identified. The PTHLH mRNA levels were measured using quantitative real-time polymerase chain reaction, while the PTHrP (parathyroid hormone-related protein) expression levels were measured using Western blotting and enzyme-linked immunosorbent assay. The interaction between TCF4 (Transcription Factor 4) and TWIST1 and the binding of the TCF4–TWIST1 complex to the PTHLH promoter were analysed using co-immunoprecipitation and chromatin immunoprecipitation. The results of the mammalian two-hybrid luciferase assay revealed that emodin inhibited TCF4–TWIST1 interaction. The effects of Polygonum cuspidatum extract (Pc-Ex), which contains emodin, on cachexia were investigated in vivo using A549 tumour-bearing mice. Ectopic expression of TCF4 upregulated PTHLH expression. Conversely, TCF4 knockdown downregulated PTHLH expression in lung cancer cells. The expression of PTHLH was upregulated in cells ectopically co-expressing TCF4 and TWIST1 when compared with that in cells expressing TCF4 or TWIST1 alone. Emodin inhibited the interaction between TCF4 and TWIST1 and consequently suppressed the TCF4/TWIST1 complex-induced upregulated mRNA and protein levels of PTHLH and PTHrP. Meanwhile, emodin-containing Pc-Ex significantly alleviated skeletal muscle atrophy and downregulated fat browning-related genes in A549 tumour-bearing mice. Emodin-containing Pc-Ex exerted therapeutic effects on lung cancer-associated cachexia by inhibiting TCF4/TWIST1 complex-induced PTHrP expression.

Physiological adaptations in early-lactation cows result in differential responses to calcium perturbation relative to nonlactating, nonpregnant cows

The peripartal cow experiences a rapid change in calcium metabolism at the onset of lactation. Research has focused on understanding how mammary-derived factors, such as serotonin (5HT) and parathyroid hormone like hormone (PTHLH), aid in coordinating these calcemic adaptations to lactation. Therefore, the aim of our study was to determine how induced subclinical hypocalcemia influences physiological responses, specifically the 5HT-PTHLH-Ca axis, in lactating and nonlactating dairy cows to elucidate the potential contribution of the mammary gland. Twelve nonlactating, nonpregnant (NL) multiparous Holstein cows and 12 early-lactation (EL) multiparous Holstein cows received either (1) a continuous 24-h intravenous solution of 0.9% NaCl or (2) 5% ethylene glycol tetraacetic acid (EGTA) solution in 0.9% NaCl (n = 6 EL, n = 6 NL per treatment) with the aim of maintaining blood ionized calcium (iCa) less than 1.0 mM. Mammary gland biopsies were taken immediately after and 48 h after termination of infusion. Blood was sampled hourly during infusion and 4, 8, 12, 24, 48, and 72 h after termination of infusion. Infusion of EGTA successfully decreased blood iCa concentrations. However, EL EGTA-infused cows required increased rates of EGTA infusion to maintain iCa below 1.0 mM. Circulating and mammary serotonin concentrations were increased in EL relative to NL cows, with no difference as a result of EGTA infusion. Mammary PTHLH expression was increased in EL cows, with highest expression observed in EL EGTA-infused cows. Collectively, these data demonstrate the robust adaptations EL cows have to maintain Ca homeostasis and the supporting roles 5HT and PTHLH may play.

PTHLH Predicts the Prognosis of Patients with Oral Leukoplakia.

BackgroundOral leukoplakia is the most common oral mucosal disease. A proportion of such cases can progress to oral squamous cell carcinoma (OSCC). The mechanism of oral leukoplakia malignant transformation is still unclear. In this study, we analyzed the expression of parathyroid hormone-like hormone (PTHLH) in oral leukoplakia and the effect on prognosis, so as to find reliable molecular markers that can predict oral leukoplakia malignant transformation.MethodsWe measured PTHrP which is coded by PTHLH in oral leukoplakia tissues of 79 cases (30 cases progressed to OSCC and 49 did not) and analyzed the clinical outcomes. Then, PTHLH expression was reduced using lentivirus-mediated small hairpin RNA (shRNA) interference to determine the biological role of PTHLH in DOK cells.ResultsPTHrP was found to be highly expressed in 38% of tissues of oral leukoplakia. There was weak or no PTHrP expression in 25 patients, moderate expression in 24 patients, and strong in 30 patients with oral leukoplakia. The expression level was associated with the degree of atypical hyperplasia and poor prognosis. The cell proliferation, invasion, migration, cell cloning, and cell cycle were affected after reducing PTHLH expression.ConclusionOur data suggest that either PTHLH or PTHrP plays a key role in the malignant transformation of oral leukoplakia and might be a reliable biomarker for predicting the carcinogenesis of oral leukoplakia.

MicroRNA expression profiles and the potential competing endogenous RNA networks in NELL-1-induced human adipose-derived stem cell osteogenic differentiation.

Studies have indicated that Nel-like molecule-1 (NELL-1) was an osteoblast-specific cytokine and some specific microRNAs (miRNAs) could serve as competing endogenous RNA (ceRNA) to partake in osteogenic differentiation of human adipose-derived stem cells (hASCs). The aim of this study was to explore the potential functional mechanisms of recombinant human NELL-1 protein (rhNELL-1) during hASCs osteogenic differentiation. rhNELL-1 was added to osteogenic medium to activate osteogenic differentiation of hASCs. High-throughput RNA sequencing (RNA-Seq) was performed and validated by real-time quantitative polymerase chain reaction. Gene ontology functional annotation and Kyoto Encyclopedia of Genes and Genomes pathway analysis were performed to detect the functions of differentially expressed miRNAs and genes. Coding-noncoding gene co-expression network and ceRNA networks were constructed to predict the potential regulatory role of miRNAs. A total of 1010 differentially expressed miRNAs and 1762 differentially expressed messenger RNAs (mRNAs) were detected. miRNA-370-3p, bone morphogenetic protein 2 (BMP2), and parathyroid hormone like hormone (PTHLH) were differentially expressed during NELL-1-induced osteogenesis. Bioinformatic analyses demonstrated that these differentially expressed miRNAs and mRNAs enriched in Rap1 signaling pathway, PI3K-Akt signaling pathway, p53 signaling pathway, Glucagon signaling pathway, and hypoxia-inducible factor-1 signaling pathway, which were important pathways related to osteogenic differentiation. In addition, miRNA-370-3p and has-miR-485-5p were predicted to interact with circ0001543, circ0002405, and ENST00000570267 in ceRNA networks. Based on the gain or loss of functional experiments by transfection, the results showed that miR-370-3p was a key regulator in osteogenic differentiation by targeting BMP2 and disturbing the expression of PTHLH, and participated in NELL-1-stimulated osteogenesis. The present study provided the primary data and evidence for further exploration on the roles of miRNAs and ceRNAs during NELL-1-induced ossification of hASCs.

Parathyroid hormone-like hormone plays a dual role in neuroblastoma depending on PTH1R expression.

We have previously reported the expression of parathyroid hormone‐like hormone (PTHLH) in well‐differentiated, Schwannian stroma‐rich neuroblastic tumors. The aim of this study was to functionally assess the role of PTHLH and its receptor, PTH1R, in neuroblastoma. Stable knockdown of PTHLH and PTH1R was conducted in neuroblastoma cell lines to investigate the succeeding phenotype induced both in vitro and in vivo. Downregulation of PTHLH reduced MYCN expression and subsequently induced cell cycle arrest, senescence, and migration and invasion impairment in a MYCN‐amplified, TP53‐mutated neuroblastoma cell line. These phenotypes were associated with reduced tumorigenicity in a murine model. We also show that PTHLH expression is not under the control of the calcium‐sensing receptor in neuroblastoma. Conversely, its production is stimulated by epidermal growth factor receptor (EGFR). Accordingly, irreversible EGFR inhibition with canertinib abolished PTHLH expression. The oncogenic role of PTHLH appeared to be a consequence of its intracrine function, as downregulation of its receptor, PTH1R, increased anchorage‐independent growth and induced a more undifferentiated, invasive phenotype. Respectively, high PTH1R mRNA expression was found in MYCN nonamplified primary tumors and also significantly associated with other prognostic factors of good outcome. This study provides the first evidence of the dual role of PTHLH in the behavior of neuroblastomas. Moreover, the identification of EGFR as a transcriptional regulator of PTHLH in neuroblastoma provides a novel therapeutic opportunity to promote a less aggressive tumor phenotype through irreversible inhibition of EGFR tyrosine kinase activity.

MiR-182-5p overexpression inhibits chondrogenesis by down-regulating PTHLH.

Human bone marrow mesenchymal stem cells (hBM-MSC) have the ability of differentiating into chondrocytes and osteoblasts. miR-182-5p promotes osteoclastogenesis and bone metastasis by up-regulating the expression of parathyroid hormone-like hormone (PTHLH). However, the function of miR-182-5p in chondrogenesis is still unknown. Mimic or inhibitor of miR-182-5p was used to upregulate or knock-down miR-182-5p expression, respectively. We analyzed chondrogenesis by Safranin O staining and Blyscan™ Sulfated Glycosaminoglycan Assay. Immunohistochemistry, real-time PCR, and Western bolts were used to detect related makers. miR-182-5p overexpression inhibited chondrogenesis. Dual-luciferase reporter assay indicated that PTHLH was one of the target genes of miR-182-5p. Further studies showed that miR-182-5p overexpression down-regulated the expression of SOX-9 and COL2A1, but up-regulated COL1A1 and COL10A1. Consistently, miR-182-5p knock-down had the opposite effects. This effect of miR-182-5p in BM-MSCs can be rescued by PTHLH overexpression. miR-182-5p may play a negative role in chondrogenesis by down-regulating PTHLH.

Gastrin induces parathyroid hormone-like hormone expression in gastric parietal cells.

Parietal cells play a fundamental role in stomach maintenance, not only by creating a pathogen-free environment through the production of gastric acid, but also by secreting growth factors important for homeostasis of the gastric epithelium. The gastrointestinal hormone gastrin is known to be a central regulator of both parietal cell function and gastric epithelial cell proliferation and differentiation. Our previous gene expression profiling studies of mouse stomach identified parathyroid hormone-like hormone (PTHLH) as a potential gastrin-regulated gastric growth factor. Although PTHLH is commonly overexpressed in gastric tumors, its normal expression, function, and regulation in the stomach are poorly understood. In this study we used pharmacologic and genetic mouse models as well as human gastric cancer cell lines to determine the cellular localization and regulation of this growth factor by the hormone gastrin. Analysis of PthlhLacZ/+ knock-in reporter mice localized Pthlh expression to parietal cells in the gastric corpus. Regulation by gastrin was demonstrated by increased Pthlh mRNA abundance after acute gastrin treatment in wild-type mice and reduced expression in gastrin-deficient mice. PTHLH transcripts were also observed in normal human stomach as well as in human gastric cancer cell lines. Gastrin treatment of AGS-E gastric cancer cells induced a rapid and robust increase in numerous PTHLH mRNA isoforms. This induction was largely due to increased transcriptional initiation, although analysis of mRNA half-life showed that gastrin treatment also extended the half-life of PTHLH mRNA, suggesting that gastrin regulates expression by both transcriptional and posttranscriptional mechanisms.NEW & NOTEWORTHY We show that the growth factor parathyroid hormone-like hormone (PTHLH) is expressed in acid-secreting parietal cells of the mouse stomach. We define the specific PTHLH mRNA isoforms expressed in human stomach and in human gastric cancer cell lines and show that gastrin induces PTHLH expression via transcription activation and mRNA stabilization. Our findings suggest that PTHLH is a gastrin-regulated growth factor that might contribute to gastric epithelial cell homeostasis.

Parathyroid Hormone-Like Hormone is a Poor Prognosis Marker of Head and Neck Cancer and Promotes Cell Growth via RUNX2 Regulation

Parathyroid Hormone-Like Hormone (PTHLH) is an autocrine/paracrine ligand that is up-regulated in head and neck squamous cell carcinoma (HNSCC). However, the cellular function and regulatory mechanism in HNSCC remains obscure. We investigated the clinical significance of PTHLH in HNSCC patients, and verified the role of RUNX2/PTHLH axis, which is stimulated HNSCC cell growth. In patients, PTHLH is a poor prognosis marker. PTHLH expression lead to increasing the cell proliferation potential through an autocrine/paracrine role and elevating blood calcium level in Nod-SCID mice. In public HNSCC microarray cohorts, PTHLH is found to be co-expressed with RUNX2. Physiologically, PTHLH is regulated by RUNX2 and also acting as key calcium regulator. However, elevations of calcium concentration also increased the RUNX2 expression. PTHLH, calcium, and RUNX2 form a positive feedback loop in HNSCC. Furthermore, ectopic RUNX2 expression also increased PTHLH expression and promoted proliferation potential through PTHLH expression. Using cDNA microarray analysis, we found PTHLH also stimulated expression of cell cycle regulators, namely CCNA2, CCNE2, and CDC25A in HNSCC cells, and these genes are also up-regulated in HNSCC patients. In summary, our results reveal that PTHLH expression is a poor prognosis marker in HNSCC patients, and RUNX2-PTHLH axis contributes to HNSCC tumor growth.

Tumor signatures of PTHLH overexpression, high serum calcium, and poor prognosis were observed exclusively in clear cell but not non clear cell renal carcinomas.

High serum calcium (Ca) due to aberrant secretion of tumor parathyroid hormone-like hormone (PTHLH) is a well-known paraneoplastic sign and is associated with poor prognosis in patients with renal cell carcinoma (RCC). However, the status of serum Ca and tumor PTHLH expression have not been verified using the 2004 World Health Organization (WHO) renal tumor classification. We retrospectively reviewed corrected serum Ca levels at initial onset (n = 683) and/or as of recurrence (n = 71) in patients with RCC. We also examined a total of 623 renal parenchymal tumor samples for PTHLH mRNA expressions by quantitative real-time PCR. High serum Ca concomitant with PTHLH overexpression in tumors was observed exclusively in clear cell RCC but not in other non clear cell subtype tumors, including papillary, chromophobe, collecting-duct, unclassified, and other rare subtype RCCs or in benign oncocytomas and angiomyolipomas. In clear cell RCC, PTHLH expression was significantly high in male patients, and was associated with a symptomatic presentation, higher grade, and higher stage cases, whereas it was not associated with VHL gene status. Univariate analyses demonstrated that high PTHLH expression was strongly associated with poor outcome both in overall survival (OS) and disease-free survival (DFS) for patients who underwent standard nephrectomy. Further multivariate Cox analyses revealed that the PTHLH expressions remained as independent prognostic parameters for OS but not for DFS. These data suggest that the previously characterized tumor signatures of high serum Ca due to high PTHLH expression and poor prognosis are clear cell RCC-specific features, whereas these characteristics are rare in non clear cell RCCs.

DLC1-dependent parathyroid hormone–like hormone inhibition suppresses breast cancer bone metastasis

Bone metastasis is a frequent complication of breast cancer that is often accelerated by TGF-β signaling; however, little is known about how the TGF-β pathway is regulated during bone metastasis. Here we report that deleted in liver cancer 1 (DLC1) is an important regulator of TGF-β responses and osteolytic metastasis of breast cancer cells. In murine models, breast cancer cells lacking DLC1 expression exhibited enhanced capabilities of bone metastasis. Knockdown of DLC1 in cancer cells promoted bone metastasis, leading to manifested osteolysis and accelerated death in mice, while DLC1 overexpression suppressed bone metastasis. Activation of Rho-ROCK signaling in the absence of DLC1 mediated SMAD3 linker region phosphorylation and TGF-β-induced expression of parathyroid hormone-like hormone (PTHLH), leading to osteoclast maturation for osteolytic colonization. Furthermore, pharmacological inhibition of Rho-ROCK effectively reduced PTHLH production and breast cancer bone metastasis in vitro and in vivo. Evaluation of clinical breast tumor samples revealed that reduced DLC1 expression was linked to elevated PTHLH expression and organ-specific metastasis to bone. Overall, our findings define a stroma-dependent paradigm of Rho signaling in cancer and implicate Rho-TGF-β crosstalk in osteolytic bone metastasis.

The Roles of Parathyroid Hormone-Like Hormone during Mouse Preimplantation Embryonic Development

Parathyroid hormone-like hormone (PTHLH) was first identified as a parathyroid hormone (PTH)-like factor responsible for humoral hypercalcemia in malignancies in the 1980s. Previous studies demonstrated that PTHLH is expressed in multiple tissues and is an important regulator of cellular and organ growth, development, migration, differentiation, and survival. However, there is a lack of data on the expression and function of PTHLH during preimplantation embryonic development. In this study, we investigated the expression characteristics and functions of PTHLH during mouse preimplantation embryonic development. The results show that Pthlh is expressed in mouse oocytes and preimplantation embryos at all developmental stages, with the highest expression at the MII stage of the oocytes and the lowest expression at the blastocyst stage of the preimplantation embryos. The siRNA-mediated depletion of Pthlh at the MII stage oocytes or the 1-cell stage embryos significantly decreased the blastocyst formation rate, while this effect could be corrected by culturing the Pthlh depleted embryos in the medium containing PTHLH protein. Moreover, expression of the pluripotency-related genes Nanog and Pou5f1 was significantly reduced in Pthlh-depleted embryos at the morula stage. Additionally, histone acetylation patterns were altered by Pthlh depletion. These results suggest that PTHLH plays important roles during mouse preimplantation embryonic development.

The Expression of PTHLH in Human Gastric Mucosa Enterochromaffin-Like Cells

BackgroundGastrectomy may disturb the body’s mineral homeostasis, with osteopenia and osteoporosis being among the late outcomes. Parathyroid hormone-like hormone (PTHLH) was detected in rat gastric enterochromaffin-like (ECL) cells in 2005, and some researchers suggested that it was the hypothetical hormone gastrocalcin that is believed to lead to osteoporosis.AimsOur objective was to learn whether PTHLH is expressed in human gastric ECL cells and to form a basic understanding about the relationship between PTHLH and gastrin.MethodsWe collected normal human gastric mucosa specimens and serum samples from 28 patients.ResultsRT-PCR and immunohistochemical analysis demonstrated weak expression of PTHLH in ECL cells at the RNA and protein levels. A low level of PTHLH expression was also found in the serum. Serum gastrin did have a significant positive correlation with the relative ratio of PTHLH mRNA to β-actin levels in gastric mucosa (r s = 0.569, p = 0.002).ConclusionsThis indicates that PTHLH has a low signal expression in human gastric ECL cells and that serum gastrin levels correlate with PTHLH RNA levels in gastric mucosa. Further work is needed to evaluate the functional role of PTHLH in ECL cells and to determine whether PTHLH is gastrocalcin.

Ext1-Dependent Heparan Sulfate Regulates the Range of Ihh Signaling during Endochondral Ossification

Exostosin1 (Ext1) belongs to a family of glycosyltransferases necessary for the synthesis of the heparan sulfate (HS) chains of proteoglycans, which regulate signaling of several growth factors. Loss of tout velu ( ttv), the homolog of Ext1 in Drosophila, inhibits Hedgehog movement. In contrast, we show that reduced HS synthesis in mice carrying a hypomorphic mutation in Ext1 results in an elevated range of Indian hedgehog (Ihh) signaling during embryonic chondrocyte differentiation. Our data suggest a dual function for HS: First, HS is necessary to bind Hedgehog in the extracellular space. Second, HS negatively regulates the range of Hedgehog signaling in a concentration-dependent manner. Additionally, our data indicate that Ihh acts as a long-range morphogen, directly activating the expression of parathyroid hormone-like hormone. Finally, we propose that the development of exostoses in the human Hereditary Multiple Exostoses syndrome can be attributed to activation of Ihh signaling.