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L C Layman

Publications and source records attributed to L C Layman.

33 records · Page 2Linked to original sources
PubMed1997

Mutation analysis of the gonadotropin-releasing hormone receptor gene in idiopathic hypogonadotropic hypogonadism.

OBJECTIVE: To determine if GnRH receptor mutations occur in patients with idiopathic hypogonadotropic hypogonadism. DESIGN: Patients and controls were studied by molecular genetic analysis. SETTING: A tertiary medical center setting. PATIENT(S): Twenty-four patients with idiopathic hypogonadotropic hypogonadism and 20 controls. INTERVENTION(S): Deoxyribonucleic acid from all individuals was analyzed by Southern blot analysis and denaturing gradient gel electrophoresis. Genomic DNA was digested with restriction enzymes, and Southern blots and denaturing gradient gel blots were constructed. Blots were hybridized with the GnRH receptor complementary DNA probe. The DNA sequencing was performed on samples from two representative patients. MAIN OUTCOME MEASURE(S): Gonadotropin-releasing hormone receptor gene structure was ascertained by comparing fragments from autoradiographs in patients and controls. Individual nucleotides were ascertained from DNA sequencing gels. RESULT(S): No GnRH receptor gene deletions or polymorphisms were identified by Southern blot analysis. New restriction-fragment melting polymorphisms using the enzymes DpnII, RsaI, and HaeIII were identified by denaturing gradient gel blots in patients and controls. CONCLUSION(S): Gonadotropin-releasing hormone receptor gene deletions or rearrangements were not observed in our idiopathic hypogonadotropic hypogonadism patients. Denaturing gradient gel electrophoresis failed to identify single-base differences unique to patients with idiopathic hypogonadotropic hypogonadism, dramatically reducing the likelihood that point mutations of the GnRH receptor gene are present in idiopathic hypogonadotropic hypogonadism.

Adolescent↗
PubMed1997

Human chorionic gonadotrophin-beta gene sequences in women with disorders of HCG production.

Women with recurrent abortion, primary unexplained infertility, and gestational trophoblastic neoplasia (GTN) manifest disordered human chorionic gonadotrophin (HCG) secretion. Mutations in the HCG beta/luteinizing hormone (LH) beta gene complex could cause aberrant HCG production in these disorders. The purpose of this study was to determine whether HCG beta gene deletions occur in women with recurrent abortion or primary unexplained infertility, and whether HCG beta gene duplications are present in women with GTN. DNA was extracted from 10 patients with unexplained recurrent abortion, 10 patients with unexplained primary infertility, 12 patients with GTN, three partners of women with GTN, and 30 controls. Southern blots were constructed and hybridized with DNA probes for HCG beta-5 and the LH beta gene. No gene deletions were identified in patients with recurrent abortion or primary unexplained infertility. Likewise, no gene duplications were identified in women with GTN. A previously described Mbol restriction fragment length polymorphism (RFLP) was identified in both patients and controls. A new Pstl RFLP was also characterized, but was present in patients and controls. Deletion/duplication mutations in the HCG beta/LH beta gene complex do not appear to be common causes of aberrant HCG production in humans with these disorders.

Abortion, Habitual↗
PubMed1997

Relevance of molecular medicine to clinical obstetrics and gynecology.

Understanding molecular biology can improve the clinical acumen of the practicing obstetrician/gynecologist. An area of basic research now becoming clinically relevant involves the G proteins and G protein-coupled receptors. Clinicians already manipulate G protein-coupled receptors in their daily practice. Examples include the administration of oxytocin (oxytocin receptors), beta-2 tocolytic agents (beta 2-adrenergic receptors), GnRH agonists (GnRH receptors), exogenous gonadotropins (FSH and LH receptors), and bromocriptine (dopamine receptor). Clinically important disorders presenting to the obstetrician/gynecologist include some forms of precocious puberty, delayed puberty, premature ovarian failure, and pituitary adenomas which are due to mutations of G proteins and G protein-coupled receptors. The importance of these proteins is demonstrated by the fact that G protein-related genes comprise about 1 percent of the human genome. Additionally, the knowledge that some G protein gene mutations are present in the germ line, and others are somatic cell in origin (and not heritable), aids in more accurate genetic counseling to patients.

Cell Communication↗
PubMed1995

The follicle-stimulating hormone receptor gene is polymorphic in premature ovarian failure and normal controls.

OBJECTIVE: To examine the FSH receptor gene for detectable abnormalities in women with premature ovarian failure. DESIGN: Study of genomic DNA from controls and from patients with 46,XX premature ovarian failure (POF). SETTING: Clinics and laboratories of university gynecology and obstetrics departments. PATIENTS: Twenty-one women with 46,XX POF and 40 normal fertile controls. INTERVENTIONS: Deoxyribonucleic acid was analyzed in patients and controls by Southern blot analysis, polymerase chain reaction (PCR), and denaturing gradient gel electrophoresis. Southern blots were hybridized with the FSH receptor complementary DNA and other smaller DNA probes. Exons 1, 5 to 6, and 10 were amplified by PCR and electrophoresed on agarose gels. Polymerase chain reaction products from exons 1 and 10 were electrophoresed on denaturing gradient gels. MAIN OUTCOME MEASURES: Fragments obtained by Southern blot analysis and PCR were compared in patients and controls. Polymerase chain reaction fragments electrophoresed on denaturing gels also were compared in patients and controls. CONCLUSIONS: No FSH receptor gene deletions or other mutations were identified in women with POF. Southern blots containing PstI- and HindIII-digested DNA revealed restriction fragment length polymorphisms in both patients and controls. Denaturing gradient gel electrophoresis analysis of PCR fragments of exon 10 also demonstrated DNA sequence polymorphisms in both patients and controls. Follicle-stimulating hormone receptor gene deletions are not common in women with POF, although the gene is polymorphic. We cannot exclude point mutations in other regions of the FSH receptor gene in some patients with POF.

Adolescent↗
PubMed1995

Molecular biology in reproductive endocrinology.

Through the study of naturally occurring mutations in humans, the creation of mutations by site-directed mutagenesis, and the production of transgenic knockout mice, further understanding of molecular reproductive endocrinology had been achieved. Mutations in the aromatase gene in females have confirmed that its deficiency results in a previously unrecognized form of sexual ambiguity with a 46,XX karyotype, and delayed puberty with multicystic ovaries. It has long been known that estrogen is necessary for skeletal growth and epiphyseal closure in the female, but aromatase and estrogen receptor gene mutations in men have demonstrated for the first time, that estrogen is important for epiphyseal closure in the male. Mutations in the steroidogenic acute regulatory protein have been recently described that demonstrate the cause for lipoid congenital adrenal hyperplasia, a disorder characterized by the complete lack of steroid production. New gene mutations in gonadotropin beta-subunits, pituitary hormones, G-protein coupled receptors, G-proteins, steroid enzymes and their receptors have also been characterized recently. Site-directed mutagenesis experiments and transgenic knockout mice have been increasingly used to study the effects of normal endocrine function. Normal functions of steroid receptor genes (steroidogenic factor-1, estrogen receptor, progesterone receptor) the glycoprotein alpha-subunit, luteinizing hormone beta, and proto-oncogenes such as RET have been better characterized by creating knockout models. Molecular biology techniques permit these types of studies which may be difficult, if not impossible, to perform otherwise in physiologic settings.

Adrenal Cortex Hormones↗
PubMed1994

Ultrasound prediction of follicle volume: is the mean diameter reflective?

OBJECTIVE: To evaluate the relationship between 2 dimensional sonographic measurement of ovarian follicles and their actual volume. DESIGN: Prospective clinical study. SETTING: The in vitro fertilization (IVF) program of a University based, tertiary care hospital. PATIENTS AND INTERVENTIONS: Sonographic categorization by shape, and measurement of 96 individual ovarian follicles immediately prior to aspiration for IVF. Each follicle was aspirated under direct ultrasound guidance and the volume recorded. The 96 follicles were visualized in a total of 14 patients from whom 2 to 27 oocytes were obtained. MAIN OUTCOME MEASURE: Total volume of each follicle. RESULTS: Round and polygonal follicles exhibited a highly significant relationship between sonographically measured mean diameter and total follicle volume. The volume of follicles that were categorized as ellipsoid was not predicted by measurement of the longest diameter, shortest diameter or mean diameter. CONCLUSION: The mean diameter of round and polygonal follicles accurately predicts total follicular volume. However, clinical decisions in ovulation induction should be modified when the follicle shape is predominantly ellipsoid because the traditionally held belief that the sonographic measurement of the follicular diameter correlates with the follicular volume does not apply in those circumstances.

Female↗
PubMed1993

Follicle-stimulating hormone beta gene structure in premature ovarian failure.

OBJECTIVE: To determine whether mutations in the gene for FSH beta are present, and possibly etiologic, in some patients with 46,XX premature ovarian failure (POF). DESIGN: DNA samples obtained from 18 study patients with POF and two menopausal fertile controls were studied by Southern blot analysis. DNA sequencing was performed in one patient. SETTING: Patients were seen in a reproductive endocrinology clinic and studied in a medical school laboratory setting at the Medical College of Georgia and Tufts University. MAIN OUTCOME MEASURES: Restriction fragment sizes on autoradiographs were compared between the study group and controls. DNA sequencing radiographs were compared between one study patient and five controls. RESULTS: Fragment sizes obtained with the restriction enzymes EcoRI, DraI, HincII, PstI, KpnI, BglI, BamHI, and BglII were similar size in both study subjects and controls using the probes pFSH beta-1.4 and pFSH beta-1.2. A previously described HindIII polymorphism was present using pFSH beta-1.2, but HindIII fragment sizes were identical in patients with ovarian failure and controls using pFSH beta-1.4. DNA sequencing of the FSH beta gene in one patient was normal. CONCLUSIONS: No mutations in the gene for FSH beta were identified in women with POF. DNA sequencing of the exons and promoter region of the FSH beta gene in one woman with POF was normal. This does not entirely exclude the possibility that smaller deletions, insertions, or point mutations of the FSH beta could be etiologic in some women with POF. The HindIII polymorphism does not appear to segregate with 46,XX POF.

Adult↗
PubMed1992

Absence of the testicular determining factor gene SRY in XX true hermaphrodites and presence of this locus in most subjects with gonadal dysgenesis caused by Y aneuploidy.

OBJECTIVES: The purpose of our study was to discover whether the testicular determining factor gene SRY (sex-determining region on Y) is present or absent in XX true hermaphrodites and in subjects with gonadal dysgenesis caused by Y aneuploidy. STUDY DESIGN: We screened five XX true hermaphrodites and 24 subjects with gonadal dysgenesis caused by Y aneuploidy for the presence or absence of SRY. With the polymerase chain reaction technique, the sequence coding the 80 amino acid-conserved motif was amplified. The 0.9 kb Hincll pY53.3 subclone, which covers the open reading frame of SRY, serves as a probe for Southern blot analysis. RESULTS: Test results for all five XX true hermaphrodites were negative for SRY. Conversely, 22 of the 24 individuals with 45,X/46,XY gonadal dysgenesis were positive for SRY, including the 10 subjects with only bilateral streak gonads. CONCLUSIONS: The absence of SRY in XX true hemaphrodites and the presence of SRY in 10 subjects with 45,X/46,XY constitution who harbored only bilateral streak gonads seem to indicate that multiple genes are involved in gonadal differentiation.

Aneuploidy↗
PubMed1992

Etiologic factors of recurrent abortion and subsequent reproductive performance of couples: have we made any progress in the past 10 years?

OBJECTIVES: We hypothesize that the diagnostic yield and pregnancy outcomes of patients with recurrent abortion have improved over the past 10 years. STUDY DESIGN: The study was performed in an academic medical center. Diagnoses and outcomes for group A, a published series of 100 patients investigated for recurrent abortion in the section between 1968 and 1977, was compared with those for group B, the 131 patients seen between 1987 and 1991. A standardized protocol was followed, enhanced by new techniques and autoimmune investigations in the latter group. Results were compiled retrospectively. Descriptive statistics and chi 2 analysis were used. RESULTS: No cause could be found in 37% of patients in group A compared with 24% of couples in group B (p less than 0.05). No clear difference could be shown in the subsequent outcomes of pregnancies. CONCLUSIONS: Our ability to establish a cause of recurrent abortion has improved slightly over the past 15 years. The gain is not yet reflected in successful pregnancy rates. Multicenter trials are indicated.

Abortion, Habitual↗
PubMed1992

Basic concepts of molecular biology as applied to pediatric and adolescent gynecology.

Recombinant DNA techniques have increased our understanding of genetic disorders and facilitated their diagnosis and may eventually lead to therapy. Expression may be assessed and quantified by Northern blot analysis, whereas in situ hybridization may localize the transcript. Southern blotting has been the standard method to identify gene mutations and RFLPs, which are useful in the diagnosis of many different disorders. Special multiallelic RFLPs produce a DNA fingerprint particularly useful in paternity testing and forensic medicine. PCR has greatly reduced the time and expense to diagnose genetic disorders, and has an enormous number of applications both clinically and at the research level. These techniques along with DNA sequencing are currently being used in the human genome project, which may some day allow "DNA typing" of individuals for any gene of interest. There is no doubt that new techniques in molecular biology will continue to evolve so that the goal of gene therapy for many disorders may be possible in the future.

Adolescent↗
PubMed1992

Identification of restriction-fragment length polymorphisms for the human chorionic gonadotropin-beta/luteinizing hormone-beta gene cluster.

OBJECTIVE: To determine whether restriction fragment length polymorphisms are present using a deoxyribonucleic acid (DNA) probe for human luteinizing hormone beta subunit (hLH-beta). If the gene for hLH-beta is polymorphic, genetic diagnosis of disorders of luteinizing hormone (hLH) and human chorionic gonadotropin (hCG) production could become possible. DESIGN: Study of genomic DNA from controls with a variety of restriction enzymes to identify polymorphisms. SETTING: Laboratories of the Department of Obstetrics and Gynecology, Department of Oral Biology, Medical College of Georgia, Augusta, Georgia. PATIENTS: Unrelated control men and women seen in clinics at the Medical College of Georgia. INTERVENTIONS: Genomic DNA was extracted from patients and digested with eight different restriction enzymes for the study of the hLH-beta gene by Southern analysis. MAIN OUTCOME MEASURE: Fragment (band) sizes on radiographs from Southern blots were compared with those from molecular weight standards. CONCLUSIONS: Restriction fragment length polymorphisms were identified for four of the restriction enzymes, DraI, HincII, MboI, and KpnI. These polymorphisms may be useful in the diagnosis of disorders of hLH and hCG production.

Chorionic Gonadotropin↗
PubMed1992

Gonadotropin-releasing hormone-associated peptide gene sequences in women with hyperprolactinemia.

OBJECTIVE: To determine if mutations in the structural gene for gonadotropin-releasing hormone (GnRH)-associated peptide are present in women with hyperprolactinemia. DESIGN: Patients with hyperprolactinemia and controls were studied retrospectively for GnRH-associated peptide gene mutations. SETTINGS: Patients seen in a clinical setting were studied at a medical school laboratory setting. PATIENTS: Fifteen women with hyperprolactinemia and two fertile controls with normal prolactin levels were studied. INTERVENTIONS: Genomic deoxyribonucleic acid (DNA) was extracted from each patient and subjected to Southern blot analysis and polymerase chain reaction (PCR). For Southern blot analysis, DNA was digested with EcoRI, XbaI, BglII, PstI, and BamHI and hybridized to two DNA probes for GnRH-associated peptide. Exons II to IV, which encode for the structural gene, were amplified by PCR. MAIN OUTCOME MEASURES: Fragment sizes from autoradiographs were compared among patients and controls. Amplified PCR products of exons II to IV of the GnRH-associated peptide were also compared. RESULTS: No large deletions, insertions, or polymorphisms were identified in women with hyperprolactinemia or controls by Southern blotting. Each of the exons was present and of normal size by PCR in the study patients and controls. CONCLUSIONS: No large deletions of the GnRH-associated peptide gene appear to be present in our patients with hyperprolactinemia. Small deletions, insertions, or point mutations are not excluded by this analysis.

Adolescent↗
PubMed1992

Gonadotropin-releasing hormone, follicle-stimulating hormone beta, luteinizing hormone beta gene structure in idiopathic hypogonadotropic hypogonadism.

OBJECTIVE: To determine if the genes for gonadotropin-releasing hormone (GnRH), follicle-stimulating hormone beta (FSH beta), and luteinizing hormone beta (LH beta) are present, and if so, whether gene structure is normal in patients with idiopathic hypogonadotropic hypogonadism (IHH). DESIGN: Patients with clinical and laboratory characteristics of IHH were studied at the deoxyribonucleic acid (DNA) level to assess gene structure. SETTING: This study took place in an academic setting. PATIENTS: Human volunteers with documented IHH and fertile controls were studied. INTERVENTIONS: Genomic DNAs were extracted from each patient, Southern blots were constructed and hybridized to DNA probes for GnRH, FSH beta, and LH beta. DNA samples were also subjected to polymerase chain reaction analysis. MAIN OUTCOME MEASURES: Gene structure was assessed by analysis of autoradiographs and gel electrophoresis of polymerase chain reaction products in both the study patients and controls. RESULTS: Each analysis for FSH beta, LH beta, and GnRH demonstrated the same sized fragments in both the study group and control group. A 1.2-kilobase fragment containing the coding region for GnRH was present in all patients with IHH and controls by polymerase chain reaction. CONCLUSIONS: The genes for GnRH, LH beta, and FSH beta are present in patients with IHH. No large deletions or rearrangements of any of these genes were identified in any of these patients.

Base Sequence↗
PubMed1990

The detection of Hind III restriction-fragment length polymorphisms using a deoxyribonucleic acid probe for the beta subunit of follicle-stimulating hormone.

Molecular diagnosis of disorders of follicle-stimulating hormone (FSH) production may become possible now that the gene for FSH beta has been characterized. Restriction-fragment length polymorphism (RFLP) analysis provides a means of organized search for molecular variants of FSH. The purpose of this study was to screen controls for the presence of RFLPs using the deoxyribonucleic acid (DNA) probe pFSH beta -1.4. Genomic DNA was digested with 12 different restriction endonucleases; Southern blots were constructed and hybridized to pFSH beta -1.4. No polymorphisms were identified with 11 enzymes. Three of 24 (12.6%) Hind III digests demonstrated a polymorphic fragment of either 5.2, 4.7, or 4.3 kb. These are the first RFLPs identified for the FSH beta gene with pFSH beta -1.4. RFLPs for FSH beta constitute the first step in the molecular analysis of disorders of FSH production.

Blotting, Southern↗