Biomedical subjects
Publications and source records attributed to R V Lloyd.
Paradoxical vocal fold movement leads to marked inspiratory stridor with adduction of the vocal folds on inspiration. A patient is presented who illustrates some of the difficulties in diagnosis and management. A range of treatments were tried including tracheostomy, laser arytenoidectomy and vocal fold lateralization.
Robson stage I or II renal carcinomas have a heterogenous clinical outcome. A variety of morphologic features and other parameters have been proposed as prognostically useful. The authors measured the DNA content and PCNA expression of 47 stage I or II renal carcinomas, and assessed the association of these measures with pathologic stage, nuclear grade, and clinical course. Approximately 56% of stage I neoplasms and 40% of stage II neoplasms were diploid. Five of 9 neoplasms in which multiple samples were analyzed manifested both aneuploid and diploid regions. PCNA expression was noted in 20 of 32 stage I neoplasms and 9 of 15 stage II neoplasms, and varied greatly among the neoplasms. Neither ploidy nor PCNA expression is associated with clinical behavior in these data. These results are different from some of those previously reported by others. These discrepancies are likely to be due to differences in methodology and the fact that there were only eight cases of metastatic disease. No single parameter will serve as a completely accurate prognostic indicator. Most individuals with these neoplasms will do well because all of the tumor has been excised.
The chromogranin/secretogranin (Cg/Sg) molecules are a family of acidic proteins present in neuroendocrine cells and tumors with secretory granules. They have been frequently used to characterize neuroendocrine cells and tumors by immunohistochemical analyses. Immunoreactivity for CgA is related to the presence of secretory granules in these tumors, so immunohistochemical staining for CgA may be absent in neuroendocrine tumors with only a few secretory granules. RNA in situ hybridization with a series of oligonucleotide probes for CgA and CgB was used to detect the mRNA transcripts for CgA and CgB with digoxigenin-labeled probes in 31 neuroendocrine tumors. These results were compared to ISH with 35S-labeled probes and with immunohistochemical staining for CgA and synaptophysin in the same neoplasms. ISH with 35S-labeled probes for CgA and B detected mRNA transcripts in 31 of 31 tumors, whereas the digoxigenin-labeled probe cocktails for CgA and B were positive in 19 of 31 cases when used separately and in 24 of 31 cases when used together. Immunohistochemical staining for CgA was positive in 22 of 31 cases and for synaptophysin in 23 of 31 cases. The CgA and B oligonucleotide probe cocktails were highly specific, since nonneuroendocrine cells and tumors did not stain and the hybridization signal was abolished by ribonuclease A pretreatment. These results indicate that non-isotopic ISH with digoxigenin-labeled probe cocktails for CgA and B or with 35S-labeled probes can be used in characterizing neuroendocrine cells and tumors in formalin-fixed paraffin-embedded tissue sections even when the CgA protein is not detected by immunohistochemistry.
Ames dwarf mice have a recessive defect that results in affected mice (df/df) with extremely hypocellular anterior pituitaries that generally lack somatotropes, lactotropes, and thyrotropes. We report detection of rare foci of GH+ cells by immunocytochemistry, suggesting that some commitment to a somatotrope cell fate can occur in the pituitaries of df/df mice. A role for GHRF in regulating somatotrope proliferation is well documented. It has been shown that expression of human GHRF (hGHRF) in transgenic mice resulted in increased somatic growth and somatotrope hyperplasia over nontransgenic littermates. To assess whether overexpression of GHRF during ontogeny might elicit a physiological response in df/df mice, we generated df/df mice expressing the hGhrf transgene. Although the somatic growth of transgenic df heterozygotes was dramatically increased over that of nontransgenic littermates, df/df mice were refractory to excess GHRF. No GHRF receptor (Grfr) transcripts were detectable in df/df fetuses or adult mice by in situ hybridization analysis. In contrast, Grfr expression is detected by e16.5 in df/+ mice. The lack of Grfr expression in df/df fetuses may account for their lack of response to GHRF and implies that the df gene product is required before e16.5.
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Nitric oxide (NO) is generated by the NO synthase family of isozymes, which is present in many mammalian cells. The constitutive NO synthase isozymes generate NO, which acts via signal transduction mechanisms in the regulation of many functions including vascular tone and blood pressure, and the inducible isozymes mediate immunological mechanisms by cytotoxic and cytostatic effects. To determine whether NO has a role in anterior pituitary cell function, immunohistochemistry and in situ hybridization analyses were used to study NO synthase expression in normal and neoplastic human pituitary tissues. Brain NO synthase was localized in the anterior pituitary in secretory and in folliculo-stellate cells and in the posterior pituitary. Pituitary adenomas had higher levels of brain NO synthase protein and mRNA compared with normal pituitaries. Endothelial NO synthase was also present in anterior and posterior pituitary cells and in endothelial cells of the pituitary. Immunoblotting studies with brain NO synthase antibodies detected a slowly migrating approximately 155-kd band and more rapidly migrating approximately 90-kd and approximately 60-kd bands. Endothelial NO synthase, but not macrophage NO synthase, was also detected in the pituitary by immunoblotting studies, confirming the immunohistochemical observations. These findings indicate that NO synthase is expressed in normal and neoplastic human pituitary tissues with increased levels of brain NO synthase protein and mRNA in adenomas compared with non-neoplastic pituitary cells and suggest that NO may play a regulatory role in hormone secretion in anterior pituitary cells.
The implantation of young children is widely accepted as a means of rehabilitating profoundly/totally deaf children. This paper will review the surgical aspects of implanting young children based on the first 50 children implanted in the Nottingham paediatric cochlear implant programme. Having thoroughly counselled the children's parents, the child is operated on under prophylactic antibiotic cover. The incision (extended endaural) is made directly down to bone and a full-thickness flap is elevated. Meticulous attention is paid to haemostasis. The steps involved in posterior tympanotomy, cochleostomy and implant insertion, often in the presence of osteogenesis, will be described. Electrophysiological testing is done systematically peroperatively. Surgical complications were few. There was no cases of facial weakness, haematoma, implant extrusion or infection. One patient, whose implant was laid in an extracochlear gutter, developed pain on electrical stimulation and was explanted. Another patient developed a retraction pocket cholesteatoma. These few complications are far outweighed by the overwhelming benefits accrued from implantation.
We studied open-lung biopsies from 17 patients with pulmonary lymphomatoid granulomatosis (LYG) using paraffin-section immunostains and Epstein-Barr virus (EBV) RNA in situ hybridization to assess the phenotype of these unique tumors and to clarify the role of EBV infection. Histologically, all cases demonstrated the characteristic mixed mononuclear cell infiltrate of lymphomatoid granulomatosis with variable numbers of cytologically atypical large lymphoid cells in a background of small lymphocytes. Paraffin-section immunostains in all cases showed a predominance of T lymphocytes. A minor population of CD20-positive large B lymphocytes was identified in 11 cases; immunoglobulin light-chain restriction was demonstrated in four of these and immunoglobulin gene rearrangements in another case. Nuclear labelling for EBV RNA was detected in 10 of these 11 cases and was confined to the population of large B lymphocytes. Staining for CD20 was absent in the remaining six cases, as was nuclear labeling for EBV RNA. However, the large atypical lymphoid cells stained for T-cell-lineage-specific antibodies in three of these cases. We conclude that some cases of lymphomatoid granulomatosis are B-cell lymphoma associated with EBV infection, whereas others are of T-cell origin and are probably unrelated to EBV infection.
Several members of the chromogranin/secretogranin (Cg/Sg) family are post-translationally processed in neuroendocrine cells and tumors to smaller peptides, some of which are biologically active. For example, CgA is processed to pancreastatin, parastatin, and other peptides. We analyzed the distribution of pancreastatin and CgA proteins in normal and neoplastic pituitaries as well as the prohormone convertases PC2 and PC3/1 (PC3), the putative processing enzymes for the Cg/Sg family, in 35 pituitary adenomas and 4 non-neoplastic pituitaries by immunohistochemistry and immunoblotting with highly specific antisera. CgA and CgB mRNAs were also examined. Pancreastatin was present in all subtypes of pituitary tumors, although prolactin-secreting adenomas expressed this peptide less frequently than did other tumor types. CgA protein and CgA mRNA expression were also restricted in prolactin adenomas and in normal prolactin cells, as shown by combined in situ hybridization and immunostaining. The prohormone convertases PC2 and PC3 were present in pituitary tumors and in non-neoplastic pituitaries. Immunoblot analysis and immunostaining showed a principal approximately 69-kd PC3 band and a approximately 68-kd PC2 band. Adrenocorticotrophic hormone-secreting adenomas expressed mainly PC3 as determined by immunoblotting and immunohistochemistry, whereas all other adenoma groups expressed predominantly PC2. These results indicate that the enzymes capable of processing CgA and other members of the Cg/Sg family to peptides with biological activity such as pancreastatin are widely expressed in human pituitary adenomas and in non-neoplastic pituitaries, with adrenocorticotrophic hormone tumors expressing predominantly PC3 and other adenomas expressing mainly PC2. The infrequent expression of CgA protein and pancreastatin peptides in normal and neoplastic prolactin cells suggests a unique role of CgA in these tumors.
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Although the relationship between pathologic features and clinical outcome is well established in adult adrenocortical neoplasms, the prognostic value of these features in pediatric adrenocortical neoplasms (PACN) is unclear. In a series of PACNs from 54 Brazilian children, the authors retrospectively investigated the prognostic value of histologic classification, ploidy, proliferative index, and size (as tumor weight or greater diameter). Histologic classification was most predictive of clinical behavior: there were no failures in 11 adenomas, 5 failures in 27 low-grade carcinomas, and 9 failures in 16 high-grade carcinomas (P = .0003). Tumor weight was predictive of failure in tumors weighing > or = 100 versus < 100 g (P = .04), and a trend was found toward failure among tumors measuring > or = 5 cm, as opposed to those < 5 cm (P = .07). Proliferative index was marginally related to failure (P = .05 at < 11% vs. > or = 11% and .07 at < 10% vs. > or = 10%), and ploidy was not significantly predictive of outcome (P = .25). Histologic type and tumor weight were the most reliable predictors of outcome in PACN.
Twenty-five pituitary adenomas were analyzed for expression of various chromogranin/secretogranin (Cg/Sg) messenger RNA (mRNA) transcripts by in situ hybridization (ISH). An additional five adenomas were also analyzed by Northern hybridization. Immunohistochemical staining for CgA and for SgIV (with monoclonal antibody HISL-19) was also performed. Most prolactin and adrenocorticotropin adenomas did not express CgA mRNA or protein, whereas growth hormone (GH) tumors had low to moderate amounts of CgA mRNA by Northern and in situ hybridization analyses and were focally positive for CgA protein. CgB, SgII, SgIII, and SgV mRNA transcripts were present in most adenomas, and SgIV protein was detected in all groups of tumors. A GH and a null cell adenoma cultured for 7 days also expressed CgA/Sg mRNA transcripts and protein. Paraffin sections of some adenomas that were negative for CgA protein had detectable CgA mRNA by in situ hybridization analysis. These results indicate that CgA mRNA and protein are more commonly expressed in glycoprotein hormone-producing tumors compared with other types of pituitary adenomas and that ISH for CgA may detect the mRNA transcripts for CgA even when CgA protein is not detected by immunohistochemistry.
Estrogen receptor (ER) was demonstrated in nontumorous and adenomatous human pituitaries by autoradiography and biochemical assays. In the present study, we investigated ER mRNA by in situ hybridization applied on paraffin section of 9 nontumorous pituitaries obtained at surgery or autopsy and 109 surgically removed adenomas. In nontumorous pituitaries, in situ hybridization combined with immunocytochemistry revealed hybridization signal in GH-, PRL-, ACTH-, TSH-, and LH/FSH-immunoreactive cells, with the highest intensity in PRL-immunoreactive cells. ER mRNA was also localized in Crooke's cells, corticotrophs extending to posterior lobe, cells lining the pars intermedia cavities, and squamous nests of pars tuberalis. The neurohypophysis, endothelium, and connective tissue expressed no ER gene. ER mRNA was present in all adenoma types, including somatotroph, lactotroph, mixed somatotroph-lactotroph, mammosomatotroph, acidophil stem cell, functioning and silent corticotroph, thyrotroph, gonadotroph, null cell adenomas, and oncocytomas. The strongest signal was seen in some lactotroph and mammosomatotroph adenomas. In 9 lactotroph adenomas exposed to bromocriptine (long-acting repeatable injectable form), the hybridization signal was weak or absent, suggesting that suppression of ER gene plays a role in the inhibition of PRL synthesis and tumor growth.
The pituitary hormones LH, FSH, and TSH are heterodimers composed of a common alpha-subunit and unique beta-subunits. We demonstrate that 4.6, 2.7, 1.49 or 0.48 kilobases (kb) mouse alpha-subunit 5'-flanking sequences are sufficient for transgene expression in both gonadotropes and thyrotropes but not in inappropriate pituitary cells. In contrast, transgenes with bovine or human alpha-subunit flanking sequences have been shown to confer reporter gene expression only to gonadotrope cells, suggesting that the elements regulating cell-specific expression may differ between species. Equal levels of reporter gene expression were conferred by 5.0 and 0.48 kb in transiently transfected thyrotrope tumor-derived cells. In contrast, in transgenic mice, high level expression was only obtained with 4.6 kb 5'-flanking sequences, indicating the presence of an enhancer element between 4.6 and 2.7 kb. The 4.6 kb of 5'-flanking sequences are sufficient for both hormonal and developmental regulation of transgene expression. Mice rendered hypothyroid by radiothyroidectomy had significantly higher levels of transgene expression than either hyperthyroid or euthyroid animals. The temporal and spatial pattern of transgene expression in Rathke's pouch paralleled that of the endogenous gene; the onset of transgene expression occurred by embryonic day 9.5. Low level expression of both the transgene and the endogenous alpha-subunit gene were detected in some unexpected peripheral sites, such as the embryonic extraocular and olfactory regions, suggesting that alpha-subunit may have a more diverse role in development than previously considered.
Craniopharyngiomas are histologically benign epithelial neoplasms of the sellar region that frequently exhibit invasive and aggressive local growth. In this study, we have investigated the presence and cellular distribution of estrogen receptor messenger ribonucleic acid by in situ hybridization in 23 surgically removed craniopharyngiomas. All craniopharyngiomas studied, including 19 adamantinomatous and 4 papillary variants, uniformly expressed the estrogen receptor gene. In all cases, an intense estrogen receptor messenger ribonucleic acid hybridization signal was demonstrated; one localized exclusively to the epithelial cells of the tumor. Connective tissue and vascular elements were devoid of hybridization signal. Coexpression of the estrogen receptor protein was also studied by immunohistochemistry. Despite the relative abundance of estrogen receptor message in all cases studied, the estrogen receptor protein was focally but conclusively detected in only two tumors. The basis of this discrepancy is unclear. Progesterone receptor protein was also studied in all cases; however, its definitive presence was noted in only one instance and, in that case, in only occasional nuclei. The expression of the estrogen receptor gene by the proliferative epithelial elements of craniopharyngiomas raises the questions of a possible hormonal component to the genesis and/or progression of the craniopharyngiomas and a potential responsiveness to therapeutic hormonal manipulation.
There have been many reports on radioisotopic in situ hybridization (ISH) studies for the demonstration of pituitary hormone mRNAs in normal pituitary gland and pituitary adenomas. Recent studies have revealed that non-radioisotopic ISH has several advantages over the radioisotopic method. Using ISH with biotinylated oligonucleotide probes, we have been able to localize various pituitary hormone mRNAs in paraffin wax or frozen sections of rat normal pituitary gland and human pituitary adenomas. For control studies we used ISH with sense probes, ISH without probes, pretreatment with ribonuclease, ISH with a probe for beta-actin and Northern blot hybridization. Using biotinylated probes, gene transcripts of rat growth hormone and prolactin were detected by Northern blot hybridization. The same biotinylated probes were used not for light microscope ISH but also for the electron microscopical demonstration of rat growth hormone and prolactin mRNAs on the polysomes of the rough endoplasmic reticula. It is emphasized that biotinylated oligonucleotide probes are useful for the analysis of pituitary endocrine function because they are applicable to the three hybridization methods, namely, Northern blot hybridization and ISH at the light and electron microscope levels.
BACKGROUND: Recent studies with the growth hormone releasing hormone (GHRH) transgenic mouse model have shown that growth hormone (GH) and prolactin (PRL) cell hyperplasia and adenomas develop in a time-dependent manner after chronic stimulation by GHRH. However, the adenomatous foci have not been shown to be neoplastic with the ability to proliferate in vitro in the absence of GHRH stimulation. EXPERIMENTAL DESIGN: A cell line was established from an enlarged pituitary from a GHRH transgenic mouse. The cells proliferated readily in culture and were characterized with respect to PRL and GH production and response to estradiol and dexamethasone. The production of chromogranin/secretogranin (Cg/Sg) mRNA transcripts and the regulation of Cg/Sg expression was also analyzed in the newly established cell line to analyze the relationship between PRL, GH, and Cg/Sg production by this cell line. RESULTS: The tumor cells responded to 10(-7) M 17 beta-estradiol (estradiol) by increasing the percentage of immunoreactive PRL-positive cells, and to dexamethasone by decreasing the percentage of PRL-positive cells and mRNA levels. Dexamethasone (10(-7) M) treatment resulted in a 3-fold reduction in PRL mRNA and CgB mRNA, whereas GH and Sg II mRNAs were both increased after dexamethasone treatment. CgA mRNA level was not changed significantly by estradiol or dexamethasone in this cell line. CONCLUSIONS: This stable transgenic cell line is regulated by estradiol and dexamethasone with changes in PRL, GH, and Cg/Sg mRNA transcript levels. There is concordant regulation of PRL and CgB mRNAs as well as GH and SgII mRNAs. These findings indicate that this cell line can be used to study the regulation and possible functions of Cg/Sg in vitro.
Many imaging methods can be used to detect pheochromocytoma, but some tumors are not detected with conventional modalities. To explore the possible usefulness of positron emission tomography (PET) after administration of 2-[fluorine-18]-fluoro-2-deoxy-D-glucose (FDG) to localize pheochromocytoma in patients with false-negative scintigrams obtained after administration of metaiodobenzylguanidine (MIBG), FDG was administered and PET was performed in two adult patients with pheochromocytomas that had never been localized despite administration of MIBG. In both patients, images were obtained dynamically for 50 minutes; then a limited truncal sequence was performed. PET enabled correct localization of the tumors. In patient 1, a tumor that had not been detected for 21 years was localized in the middle mediastinum; in patient 2, a pheochromocytoma was detected in the right adrenal gland. PET performed after administration of FDG may be useful for localization of pheochromocytomas that do not accumulate MIBG.