Pseudohypoparayhyroidism (PHP) is a rare autosomal dominant or recessive genetic disorder characterized by low calcium, high phosphorus, and target organ resistance to parathyroid. The clinical characteristics and genetic features in 4 patients with Type Ib PHP in the Third Xiangya Hospital, Central South University, have been reviewed. All 4 patients had low calcium, high phosphorus, and parathyroid resistance. Among them, 2 patients had slightly elevated thyroid stimulating hormone and mild features of Albright's hereditary osteodystrophy, and one patient had hypokalemia. No guanine nucleotide-binding protein alpha-stimulating activity polypeptide 1 (GNAS) and gene variant associated with hypokalemia were identified using the whole exome sequencing. The results of the methylation-specific multiple ligation-dependent probe amplification showed that there were abnormal methylation of the upstream differentially methylated regions of GNAS in the 4 patients. There were phenotype overlap among the various subtypes of PHP. Detection of GNAS gene methylation in patients with clinical suspicion of Type Ib PHP is helpful for the diagnosis and treatment of PHP.
Humans ; Chromogranins/genetics* ; GTP-Binding Protein alpha Subunits, Gs/genetics* ; Hypokalemia ; Calcium ; Pseudohypoparathyroidism/genetics* ; Phosphorus

OBJECTIVE: To preliminarily investigate the relationship between stimulatory G protein α subunit (GNAS) and thyroid hormone receptor α (THRA) gene mutations and clinical phenotypes in children with congenital hypothyroidism (CH). METHODS: A total of 70 children with CH diagnosed by neonatal screening were enrolled. Their peripheral blood samples were collected to extract genomic DNA. GNAS and THRA genes were screened for mutations using next-generation sequencing. Bioinformatics software was used to analyze the pathogenicity of gene mutations. RESULTS: Of the 70 children with CH, nine missense mutations (three known mutations and six novel mutations) in the GNAS gene were detected in three patients (4%), and one gene polymorphism, c.508A>G(p.I170V), in the THRA gene was detected in four patients. The analysis results of bioinformatics software and ACMG/AMP guidelines showed that the two GNAS gene mutations [c.301C>T(p.R101C) and c.334G>A(p.E112K)] were more likely to be pathogenic. Three children with GNAS gene mutations showed different degrees of hypothyroidism. CONCLUSIONS GNAS gene mutations are related to the development of CH, and children with CH have different clinical manifestations. THRA gene mutations may not be associated with CH.
Chromogranins ; genetics ; Congenital Hypothyroidism ; GTP-Binding Protein alpha Subunits, Gs ; genetics ; Genes, erbA ; Humans ; Infant, Newborn ; Mutation ; Phenotype ; Thyroid Hormone Receptors alpha ; genetics

OBJECTIVE: To explore the genetic etiology of a Chinese pedigree affected with pseudohypoparathyroidism. METHODS: Peripheral blood samples of the proband and his parents were collected and subjected to trio-whole exome sequencing (trio-WES). Candidate variants were verified among the pedigree and 50 randomly selected healthy individuals through analysis of restriction fragment length polymorphism. Short tandem repeat (STR) linkage analysis was used to verify the parental origin of the pathogenic variants. RESULTS: Trio-WES and Sanger sequencing showed that the proband and his mother had both harbored a c.121C>G (p.His41Asp) variant of the GNAS gene, which was not found in other family members and the 50 healthy controls. The variant was not found in international databases. Based on guidelines from the American College of Medical Genetics and Genomics, the variant was predicted to be likely pathogenic. CONCLUSION The novel c.121C>G variant of the GNAS gene probably underlay the disease in this pedigree. Above finding has enriched the spectrum of GNAS gene variants.
Female ; Humans ; Pedigree ; East Asian People ; Mothers ; Exome Sequencing ; Pseudohypoparathyroidism/genetics* ; Mutation ; China ; Chromogranins/genetics* ; GTP-Binding Protein alpha Subunits, Gs/genetics*

OBJECTIVETo investigate the clinical features, mutation of the GNAS1 and pathogenesis of progressive osseous heteroplasia (POH).

METHODThe typical clinical, pathological and radiographic features of a boy with POH were collected and summarized following family survey. The GNAS1 gene sequence of all family members were amplified by polymerase chain reaction (PCR) and the products were sequenced directly to identify the mutations. A literature review and long-term follow up were also conducted.

RESULTThe patient was an 11-year-old boy who had the onset in infancy, which indicates a chronic progressive cause of disease. The clinical features include the unsmooth local skin of the right shank where spread many rigid rice-like or irregular slabby uplifts, slabby bone-like sclerosis on the left lower mandible, left masticatory muscles, in lateral subcutaneous site of left hip joint and deep tissue, accompanied by gradually progressive difficulty in opening mouth. Histopathology showed that there were loosened hyperplasia of fibroblast and interstitial edema with punctiformed ossification. Radiographs showed flocculence hyperdense image in the subcutaneous tissues and muscles around left lower mandible, and the left masticatory muscles were obviously involved. The 3-dimensional computed tomography showed dislocations of the left temporomandibular joint. Sheeted hyperdense image with inequable density could be noted in lateral muscles of the left hip. And lamellar hyperdense image parallel to the long axis of the bone could be seen in the subcutaneous dorsum of the left foot and achilles tendon. Macro-thumb and of brachydactylia of the hands and feet were not present. The level of calcium, phosphorus and alkaline phosphatase in the blood were normal. Brother of same father but different mothers was free of the disease and no patient of the same disease was found in maternal line and paternal lines. A mutated allele in exon 7 and a polymorphism in exon 5 were found in GNAS1 gene in both of the patient and his father.

CONCLUSIONThere is possibility/likelihood/probability that Chinese children could develop POH. Translocated dermal ossification began in infancy and shows a progressive cause in childhood. The disease is characterized by the heterotopic ossification of the skin, deep tissue, muscles and facial surface tissues. The location of the mutation in this study was different from that reported in abroad studies although exist in the same exons.

Child ; Chromogranins ; DNA Mutational Analysis ; Exons ; GTP-Binding Protein alpha Subunits, Gs ; genetics ; Humans ; Male ; Mutation ; Ossification, Heterotopic ; diagnosis ; genetics ; pathology ; Pedigree

Serotonin receptor subtype 6 (5-HT(6)) is a neurotransmitter receptor, which is involved in various brain functions such as memory and mood. It mediates signaling via the interaction with a stimulatory G-protein. Especially, the third intracellular loop (iL3) of 5-HT(6) and the alpha subunit of stimulatory G protein (Galpha(s)) are responsible for the signaling process of 5-HT(6). Chemical compounds that could inhibit the interaction between the iL3 region of 5-HT(6) and Galpha(s) were screened from a chemical library consisted of 5,600 synthetic compounds. One of the identified compounds bound to Galpha(s) and effectively blocked the interaction between Galpha(s) and the iL3 region of 5-HT(6). The identified compound was further shown to reduce the serotonin-induced accumulation of cAMP in 293T cells transformed with 5-HT(6) cDNA. It also lowered the Ca2+ efflux induced by serotonin in cells expressing 5-HT(6) and chimeric Galpha(s5/q). These results indicate that the interaction between the iL3 of 5-HT(6) and Galpha(s) can be exploited for screening of regulatory compounds against the signaling pathway of 5-HT(6).
Animals ; Calcium/metabolism ; Cell Line ; Cephalosporins/*pharmacology ; Cricetinae ; Cricetulus ; Cyclic AMP/biosynthesis ; GTP-Binding Protein alpha Subunits, Gs/antagonists & inhibitors/*metabolism ; Humans ; Receptors, Serotonin/*drug effects/metabolism/*physiology ; Serotonin/pharmacology ; Serotonin Antagonists/pharmacology ; Signal Transduction

McCune-Albright syndrome (MAS) is a rare disorder that develops from an activating mutation in the Gs gene. It is characterized by an association with Polyostotic fibrous dysplasia, and precocious puberty, Caf-au-lait pigmentation, and other endocrinopathies that result from the hyperactivity of a variety of endocrine glands. Recently we encountered a patient with MAS with fibrous dysplasia, skin pigmentation, acromegaly, hyperprolactinemia and a thyroid nodule. A 23-year-old male presented for an evaluation of a change in his facial structures. Fibrous dysplasia was diagnosed by a bone biopsy and radiographic studies. The GH level increased paradoxically after an oral glucose load. The plasma prolactin, IGF-1 and alkaline phosphatase were high. Thyroid ultrasonography revealed multiple nodules. The brain MRI demonstrated a mass in the left pituitary gland. Genetic analysis identified a change from Arg (CGT) at codon 201 to Cys (TGT).
Thyroid Diseases/etiology/genetics ; Puberty, Precocious/etiology/genetics ; Mutation ; Male ; Hyperprolactinemia/etiology/genetics ; Humans ; GTP-Binding Protein alpha Subunits, Gs/*genetics ; Fibrous Dysplasia, Polyostotic/*diagnosis/genetics/pathology ; Cafe-au-Lait Spots/etiology/genetics ; Adult ; Acromegaly/*diagnosis/etiology

The aim of the present study is to explore the mechanism of estrogen on regulating cardiac function disorder by adjusting the stimulating adenylate cyclase G &alpha; protein (G&alpha;s)-cycle adenosine monophosphate (cAMP) signal pathway. Adult female rats were randomly divided into five groups: sham group, ovariectomized group (OVX), OVX and 17β-estradiol given group (OVX+E₂), OVX and isoprenaline injected group (OVX+ISO), OVX and 17β-estradiol, isoprenaline injected group (OVX+E₂+ISO). Rats were ovariectomized, and two weeks later, OVX+E₂group was injected with E₂, OVX+ISO group was injected with ISO, OVX+E₂+ISO group was injected with E₂and ISO. Another four weeks later, the hemodynamic parameters were monitored by carotid artery intubation: left ventricular systolic pressure (LVSP), left ventricular end-diastolic pressure (LVEDP), maximal differentials of left ventricular developed pressure (+dp/dt(max)), and minimal differentials of left ventricular developed pressure (-dp/dt(max)). Brain natriuretic peptide (BNP) and cAMP concentration in plasma were determined; G&alpha;(s) protein expression in myocardium was determined. The results showed that the hemodynamic parameters, the concentration of BNP and cAMP in plasma had no significant changes after ovariectomy compared with sham group. But after isoprenaline injection in ovariectomized rats, LVSP and +dp/dt(max) declined (P < 0.01), LVEDP and -dp/dt(max) elevated (P < 0.01); plasma BNP concentration increased (P < 0.01); plasma cAMP concentration decreased (P < 0.01), compared with OVX group. Further estrogen supplements improved the heart function treated by isoprenaline: LVSP and +dp/dt(max) elevated (P < 0.01), LVEDP and -dp/dtmax declined (P < 0.05, P < 0.01); the plasma BNP concentration decreased (P < 0.01); the plasma cAMP concentration increased (P < 0.01). Estrogen had no significant influence on G&alpha;s protein expression. The results suggest that estrogen can alleviate myocardial injury and regulate cardiac function disorder by increasing cAMP level, finally improved the excessive suppression of myocardium.
Animals ; Cyclic AMP ; blood ; Estradiol ; pharmacology ; Estrogens ; pharmacology ; Female ; GTP-Binding Protein alpha Subunits, Gs ; metabolism ; Hemodynamics ; Isoproterenol ; adverse effects ; Myocardium ; pathology ; Natriuretic Peptide, Brain ; blood ; Ovariectomy ; Rats ; Signal Transduction

OBJECTIVETo observe anti-arrhythmic effect of acupuncture pretreatment in the rat of myocardial ischemia and reperfusion (MIR) and to explore the role of cAMP and Gsa protein in beta-adrenergic receptor signaling.

METHODSMIR was produced by ligation and reperfusion of the left anterior descending coronary artery in the rat. Arrhythmic score, content of cAMP and Gsalpha protein in ischemic myocardium were compared among the normal control (NC), ischemia and reperfusion (IR), electroacupuncture (EA) and EA plus propranolol (EAP) groups.

RESULTSThe arrhythmic score in the IR group at 10 min after reperfusion was higher than the NC group (P < 0.01); in the EA group the score was decreased (P < 0.01 vs the IR group); the score in the EAP group was similar to the IR group, much higher than the EA group (P < 0.01). The similar results for the contents of cAMP and Gsalpha protein were found in the ischemic myocardium. It is suggested that EA pretreatment significantly attenuates the arrhythmic incidence rate and the enhancement of the contents of myocardial cAMP and Gsalpha protein induced by MIR, and the attenuating effect is significantly inhibited by the intraperitoneal pretreatment of propranolol, a specific beta-adrenoceptor antagonist.

CONCLUSIONPretreatment of EA can produce anti-arrhythmic effect in the rat of MIR, which is mediated by the post-receptor signaling pathway of beta-adrenergic receptor.

Acupuncture Therapy ; Animals ; Arrhythmias, Cardiac ; prevention & control ; Calcium ; metabolism ; Cyclic AMP ; analysis ; GTP-Binding Protein alpha Subunits, Gs ; analysis ; Male ; Myocardial Ischemia ; metabolism ; therapy ; Rats ; Rats, Wistar ; Receptors, Adrenergic, beta ; physiology ; Signal Transduction ; physiology

Stimulatory heterotrimeric GTP-binding proteins (Gs protein) stimulate cAMP generation in response to various signals, and modulate various cellular phenomena such as proliferation and apoptosis. This study aimed to investigate the effect of Gs proteins on gamma ray-induced apoptosis of lung cancer cells and its molecular mechanism, as an attempt to develop a new strategy to improve the therapeutic efficacy of gamma radiation. Expression of constitutively active mutant of the alpha subunit of Gs (GalphasQL) augmented gamma ray-induced apoptosis via mitochondrial dependent pathway when assessed by clonogenic assay, FACS analysis of PI stained cells, and western blot analysis of the cytoplasmic translocation of cytochrome C and the cleavage of caspase-3 and ploy(ADP-ribose) polymerase (PARP) in H1299 human lung cancer cells. GalphasQL up-regulated the Bak expression at the levels of protein and mRNA. Treatment with inhibitors of PKA (H89), SP600125 (JNK inhibitor), and a CRE-decoy blocked GalphasQL-stimulated Bak reporter luciferase activity. Expression of GalphasQL increased basal and gamma ray-induced luciferase activity of cAMP response element binding protein (CREB) and AP-1, and the binding of CREB and AP-1 to Bak promoter. Furthermore, prostaglandin E2, a Galphas activating signal, was found to augment gamma ray-induced apoptosis, which was abolished by treatment with a prostanoid receptor antagonist. These results indicate that Galphas augments gamma ray-induced apoptosis by up-regulation of Bak expression via CREB and AP-1 in H1299 lung cancer cells, suggesting that the efficacy of radiotherapy of lung cancer may be improved by modulating Gs signaling pathway.
Apoptosis/*radiation effects ; Cell Line, Tumor ; Cyclic AMP Response Element-Binding Protein/metabolism ; GTP-Binding Protein alpha Subunits, Gs/*metabolism ; *Gamma Rays ; Heterotrimeric GTP-Binding Proteins/metabolism ; Humans ; Lung/*cytology/physiology/radiation effects ; Lung Neoplasms ; Transcription Factor AP-1/metabolism ; *Up-Regulation ; bcl-2 Homologous Antagonist-Killer Protein/*metabolism

Heterotrimeric GTP-binding proteins (G proteins) transduce extracellular signals into intracellular signals by activating effector molecules including adenylate cyclases that catalyze cAMP formation, and thus regulate various cellular responses such as metabolism, proliferation, and apoptosis. cAMP signaling pathways have been reported to protect cells from ionizing radiation-induced apoptosis, but however, the protective mechanism is not clear. Therefore, this study aimed to investigate the signaling molecules and the mechanism mediating the anti-apoptotic action of cAMP signaling system in radiation-induced apoptosis. Stable expression of a constitutively active mutant of G alpha s (G alpha sQL) protected gamma ray-induced apoptosis which was assessed by analysis of the cleavages of PARP, caspase-9, and caspase-3 and cytochrome C release in SH-SY5Y human neuroblastoma cells. G alpha sQL repressed the gamma ray-induced down-regulation of Bcl-xL protein, but transfection of Bcl-xL siRNA increased the gamma ray-induced apoptosis and abolished the anti-apoptotic effect of G alpha sQL. G alpha sQL decreased the degradation rate of Bcl-xL protein, and it also restrained the decrease in Bcl-xL mRNA by increasing the stability following ionizing irradiation. Furthermore, prostaglandin E2 that activates G alpha s was found to protect gamma ray-induced apoptosis, and the protective effect was abolished by treatment with prostanoid receptor antagonist specific to EP2/4R subtype. Moreover, specific agonists for adenosine A1 receptor that inhibits cAMP signaling pathway augmented gamma ray-induced apoptosis. From this study, it is concluded that Galphas-cAMP signaling system can protect SH-SY5Y cells from gamma ray-induced apoptosis partly by restraining down-regulation of Bcl-xL expression, suggesting that radiation-induced apoptosis can be modulated by GPCR ligands to improve the efficiency of radiation therapy.
Apoptosis/*physiology/*radiation effects ; Base Sequence ; Cell Line, Tumor ; Cyclic AMP/metabolism ; DNA Primers/genetics ; Down-Regulation/radiation effects ; GTP-Binding Protein alpha Subunits, Gs/*metabolism ; Gamma Rays ; Humans ; Neuroblastoma/genetics/metabolism/pathology ; RNA, Small Interfering/genetics ; Signal Transduction ; bcl-X Protein/genetics/*metabolism