Biomedical subjects

Q Hamid

Publications and source records attributed to Q Hamid.

At least 163 records · Page 9Linked to original sources

PubMed1991

Messenger RNA expression of the cytokine gene cluster, interleukin 3 (IL-3), IL-4, IL-5, and granulocyte/macrophage colony-stimulating factor, in allergen-induced late-phase cutaneous reactions in atopic subjects.

Cryostat sections from skin biopsies from 24-h allergen-induced late-phase cutaneous reactions (LPR) in 14 human atopic subjects were hybridized with 35S-labeled RNA probes for a number of cytokines. mRNA was detected for interleukin 3 (IL-3) (8/14), IL-4 (10/14), IL-5 (11/14), and granulocyte/macrophage colony-stimulating factor (GM-CSF) (13/14). Only 5 of 14 gave hybridization signals for IL-2, and 0 of 14 for interferon gamma. Biopsies from diluent controls gave only occasional weak signals. These results suggest that cells infiltrating the site of the 24-h LPR transcribe mRNA for the IL-3, IL-4, IL-5, and GM-CSF gene cluster and support the hypothesis that atopy is associated with preferential activation of cells having a similar cytokine profile to the murine T helper type 2 subset.

Adolescent↗

PubMed1991

Oncoprotein immunoreactivity in human endocrine tumours.

Abnormally expressed oncogenes are implicated in neoplastic transformation. We have investigated a series of endocrine tumours using immunocytochemistry as a first screening tool to detect oncogene expression. Paraffin sections of 44 pulmonary small cell carcinomas, 15 pulmonary atypical carcinoids, 12 bronchial carcinoids, 28 medullary thyroid carcinomas, 27 phaeochromocytomas, and 17 insulinomas were immunostained with antibodies to c-erbB-2, c-myc, L-myc, and N-myc. Diffuse immunoreactivity was detectable for c-erbB-2 in 8 out of 44 (18 per cent) pulmonary small cell carcinomas, 3 out of 15 (20 per cent) pulmonary atypical carcinoids, and 6 out of 27 (22 per cent) phaeochromocytomas; for c-myc in 18 out of 44 (41 per cent) pulmonary small cell carcinomas and 5 out of 15 (33 per cent) pulmonary atypical carcinoids; for N-myc in 6 out of 28 (21 per cent) medullary thyroid carcinomas; and for L-myc in 4 out of 27 (15 per cent) phaeochromocytomas. There was considerable intratumoral and intertumoral heterogeneity and, in each tumour group, no relationship was found between tumour pattern, mitotic index, and oncoprotein immunoreactivity. These results suggest that oncogene products are present in a proportion of endocrine tumours, and that specific oncoproteins seem to be related to tumour type but not to other histopathological findings. Thus, oncoprotein detection may be a useful tool for identifying subsets of endocrine tumours that are not otherwise recognizable morphologically.

Endocrine System Diseases↗

PubMed1991

Expression of chromogranin A mRNA in small cell carcinoma of the lung.

Chromogranin A (CgA) is the most specific neuroendocrine marker but in small cell carcinoma of the lung (SCCL) CgA immunoreactivity cannot always be detected, possibly owing to poor granularity. The localization of CgA mRNA could be a more useful indicator of CgA synthesis in SCCL. We have therefore studied the expression of CgA mRNA in SCCL, using in situ hybridization and Northern blot analysis, comparing it with CgA immunoreactivity. The expression of CgA mRNA and immunoreactivity in other types of lung tumours was also investigated. Freshly fixed, surgically resected SCCL (n = 5), adenocarcinoma (n = 10), squamous cell carcinoma (n = 10), large cell carcinoma (n = 5), and primary lymphoma (n = 2) were examined. Weak immunoreactivity to CgA was seen in one SCCL but all five strongly expressed CgA mRNA. The specificity of the in situ hybridization signal and the probe was confirmed by the use of a sense probe and Northern blot analysis. Non-SCCLs were negative for both CgA mRNA and immunoreactivity. This study provides evidence for CgA synthesis in SCCL and confirms the specificity of CgA for neuroendocrine differentiation. In situ hybridization appears to be a very useful tool for investigating the nature of tumour cells, especially in the absence of the end product of the gene transcript.

Blotting, Northern↗

PubMed1991

Localization of beta pre-protachykinin mRNA in nodose ganglion.

The neurons which synthesize tachykinins in the capsaicin-sensitive afferent nerves of the respiratory tract are largely localized to the nodose ganglia. Using a radiolabelled antisense cRNA probe constructed from cDNA for the major precursor of substance P and neurokinin A (beta-preprotachykinin: beta-PPT), we have localized specific mRNA for this peptide in neurons of the nodose ganglion of rat using in situ hybridization. 26% of neurons gave a positive hybridization signal, which was in agreement with the same proportion of cell bodies showing substance P-like immunoreactivity. The specificity of the hybridization was confirmed by the absence of labelling using RNase pre-treatment and a sense probe having the same sequence as beta-PPT mRNA. This approach may now allow the investigation of factors which regulate synthesis of tachykinins at a gene transcriptional level.

Animals↗

PubMed1991

TNF alpha mRNA expression in allergic inflammation.

Using the technique of in situ hybridization, we have attempted to identify messenger RNA for tumour necrosis factor-alpha (TNF alpha) in cells infiltrating allergen-induced late phase reaction (LPR) of the skin and the nose of atopic subjects. We have also compared the number of TNF alpha mRNA positive cells in bronchoalveolar lavage (BAL) from atopic asthmatics and normal controls. Twenty-four hours after local allergen challenge, 12/14 skin biopsies and 9/10 nasal biopsies had positive hybridization signals for TNF alpha mRNA whereas only 4/14 and 2/10 biopsies were positive in the relevant diluent controls. Compared with diluent sites significantly increased numbers of cells expressing mRNA for TNF alpha were observed in the LPR of skin (P less than 0.004) and nose (P less than 0.006). All BAL from asthmatics (n = 10) and from normal volunteers (n = 10) had cells showing positive hybridization signals for TNF alpha mRNA but these were at increased frequency in asthmatics (P less than 0.001). These results suggest that TNF alpha may be an important cytokine in atopic allergic inflammation.

Adolescent↗

PubMed1991

Epithelial changes following bone marrow transplantation: cytological atypia and epithelial endocrine cells.

This report describes the histological and immunocytochemical findings in gastrointestinal biopsies and skin of a patient with chronic granulocytic leukaemia which progressed to blastic transformation, who was then treated with chemotherapy and total body irradiation followed by allogeneic bone marrow transplantation. The gut showed endocrine cells in the lamina propria and these had an immunophenotype similar to the glandular epithelium at the sites studied (stomach, duodenum and rectum), supporting the idea that the endocrine cells of the gut are more resistant to the effect of radiation, chemotherapy and graft-versus-host disease (GVHD) than are other cell types in the epithelium, and that lamina proprial endocrine cells are epithelially derived in this situation. Epidermis and gut epithelium also showed marked atypia due to the conditioning regimen and GVHD, and this case illustrates the possibility of misdiagnosis of carcinoma in this increasingly common situation. An additional, unusual feature of this case was the presence of a B-cell UCHL1 + ve lymphoma of the colon at autopsy, 122 days post-transplantation.

Bone Marrow Transplantation↗

PubMed1991

Interleukin-5 mRNA in mucosal bronchial biopsies from asthmatic subjects.

Using in situ hybridization, we have investigated the expression of interleukin-5 (IL-5) mRNA in bronchial biopsies from asthmatics (n = 10) and controls (n = 9). The number of IL-5-nRNA-positive cells were compared with the number of CD25+ and EG2+ cells and total eosinophil counts. Specific hybridization signals for IL-5 mRNA were demonstrated in 6 out of the 10 asthmatic subjects but in none of the controls. The 6 IL-5-mRNA-positive asthmatics tended to have more severe disease and showed a significant increase in the degree of infiltration of the bronchial mucosa by activated T lymphocytes and eosinophils.

Asthma↗

PubMed1991

Expression of mRNA for interleukin-5 in mucosal bronchial biopsies from asthma.

We have attempted to identify mRNA for IL-5 in endobronchial mucosal biopsies from asthmatics and controls, using the technique of in situ hybridization. Bronchial biopsies were obtained from 10 asthmatics and 9 nonatopic normal controls. A radio-labeled cRNA probe was prepared from an IL-5 cDNA and hybridized to permeabilized sections. These were washed extensively before processing for autoradiography. An IL-5-producing T cell clone derived from a patient with the hyper-IgE syndrome was used as a positive control. As a negative control, sections were also treated with a "sense" IL-5 probe. Specific hybridization signals for IL-5 mRNA were demonstrated within the bronchial mucosa in 6 out of the 10 asthmatic subjects. Cells exhibiting hybridization signals were located beneath the epithelial basement membrane. In contrast, there was no hybridization in the control group. No hybridization was observed with the sense probe. The six IL-5 mRNA-positive asthmatics tended to have more severe disease than the negative asthmatics, as assessed by symptoms and lung function, and showed a significant increase in the degree of infiltration of the bronchial mucosa by secreting (EG2+) eosinophils and activated (CD25+) T lymphocytes. Within the subjects who showed positive IL-5 mRNA, there was a correlation between IL-5 mRNA expression and the number of CD25+ and EG2+ cells and total eosinophil count. This study provides evidence for the cellular localization of IL-5 mRNA in the bronchial mucosa of asthmatics and supports the concept that this cytokine regulates eosinophil function in bronchial asthma.

Adult↗

PubMed1989

Non-isotopic RNA probes. Comparison between different labels and detection systems.

Several studies have shown the use of non-radioactive labelled DNA probes for in situ hybridisation, mainly to identify cellular DNA. In this study mRNA in situ hybridisation was performed on rat pituitary with biotinylated complementary (c) RNA probes for rat prolactin and growth hormone (GH), and compared with radioactive 35S-radiolabelled probes. Biotinylated cRNA probes were labelled with either biotin-11-UTP or with allylamine-UTP, the latter method being able to produce a higher yield of labelled RNA. Different detection systems were tested, and hybridisation signal was seen in cells of anterior pituitary with both types of biotinylated probes. The signals were detected using either avidin-biotin-complex with peroxidase (ABC), peroxidase-anti-peroxidase (PAP) or gold-silver methods. ABC peroxidase detected using glucose oxidase-diaminobenzidine (DAB)-nickel solution appeared to be the best method for detecting labelled RNA probes, with very strong signal and low background. The biotinylated probes were comparable in sensitivity to the radiolabelled probes in detecting prolactin and GH mRNAs in the anterior lobe of the rat pituitary. These results indicate an alternative methods of labelling and detection of biotinylated probes which could have a potential role in research and diagnostic techniques.

Allylamine↗

PubMed1988

Oligo-riboprobes. Tools for in situ hybridisation.

In situ hybridisation detection of mRNAs using riboprobes has become a widely used technique. However, the identification of cells producing closely-related yet distinct mRNAs is difficult with the usual size probes. Moreover, it is not always easy to obtain the required cDNA essential for cRNA probe synthesis. To avoid these problems, we have used synthetic oligodeoxynucleotides to generate short, single stranded RNA probes ("oligo-riboprobes"). These probes can be labelled to very high (10(9) cpm/micrograms) specific activity and can be prepared for any published nucleotide sequence. We have used these probes to localise beta (preprotachykinin) PPT mRNA producing neurons in rat hypothalamus and bowel. The results were compared to that obtained with cRNA probes generated from beta preprotachykinin cDNA.

Animals↗

PubMed1988

Localisation of mRNA and co-expression and molecular forms of GRP gene products in endocrine cells of fetal human lung.

The presence of bombesin (gastrin-releasing peptide, GRP)-like immunoreactivity in mucosal endocrine cells of human fetal lung is well established. In this study we have investigated the localisation of pro-GRP mRNA and GRP gene products and compared the distribution and levels of extractable GRP- and C-terminal flanking peptide of human pro-GRP-like immunoreactivity in order to verify synthesis and to investigate their coexistence and molecular forms. Human fetal lungs (14 to 23 weeks gestation) were immunostained, and extracts were assayed using region-specific antisera to pro-GRP. Additional antisera to chromogranin and protein gene product 9.5 (PGP 9.5) were used for immunostaining by the peroxidase anti-peroxidase technique and for double immunofluorescence staining using antisera raised in two species. Immunoreactivity for both bombesin (GRP) and flanking peptide was seen mainly in the same endocrine cells, but more cells were stained with antisera to flanking peptide than with antiserum to bombesin (GRP). In situ hybridisation showed that pro-GRP mRNA was present and thus synthesis of the peptides was taking place. Endocrine cells and nerve fibres were PGP 9.5-immunoreactive, and a subset of cells was immunoreactive for bombesin gene products. Radioimmunoassay and chromatography show that pro-GRP is present in both the uncleaved and cleaved forms, and, in agreement with immunocytochemistry results, that an excess of C-terminal peptide of pro-GRP is detectable. It is therefore concluded that GRP-like peptides and flanking peptide are co-localised in human pulmonary endocrine cells, but the latter is found in larger concentrations than free GRP. Thus GRP-like peptides may be secreted separately from the flanking peptide(s) of pro-GRP.(ABSTRACT TRUNCATED AT 250 WORDS)

Chromatography, Gel↗

PubMed1988

Combined use of in situ hybridisation and immunocytochemistry for the investigation of prolactin gene expression in immature, pubertal, pregnant, lactating and ovariectomised rats.

We have investigated the use of in situ hybridisation together with immunocytochemistry for the study of endocrine cell function, using as an example the expression of prolactin messenger RNA (mRNA) in pituitaries of rats under various endocrinological conditions. In situ hybridisation using a 32P-labelled cRNA probe for rat prolactin was carried out on sections of 4% paraformaldehyde-fixed pituitaries from prepubertal, pubertal, pregnant, lactating and ovariectomised rats and adjacent sections were immunostained for prolactin. Northern gel analysis was performed on total RNA extracts of pregnant, lactating and control pituitaries. While in ovariectomised rat pituitaries both prolactin immunoreactivity and prolactin mRNA were decreased, no differences in prolactin immunostaining were seen between prepubertal, pubertal, pregnant or lactating rats and controls, even when the supra-optimal dilution technique was used. However, using in situ hybridisation, prolactin mRNA signal was increased in prepubertal rats, and with hybridisation and northern gel analysis the signal was reduced in pregnant rats and markedly increased in lactating rats. The combined use of in situ hybridisation and immunocytochemistry provides morphological information concerning endocrine gene expression and protein synthesis in the pituitary gland.

Animals↗

PubMed1988

Simultaneous demonstrations of neuropeptide Y gene expression and peptide storage in single neurons of the human brain.

A combination of in situ hybridization for neuropeptide Y mRNA that used a 32P-labeled complementary RNA probe and immunocytochemistry with polyclonal antibodies against neuropeptide Y were applied to human cortical brain samples to simultaneously localize neuropeptide Y and its mRNA. These two techniques allowed simultaneous identification of neuropeptide Y gene expression and peptide storage in single neurons of the human brain.

Brain↗

PubMed1988

Localization of bombesin-like peptides in tumors.

The localization of bombesin gene products in neuroendocrine tumors was achieved by a number of techniques used in combination. These included immunocytochemistry, radioimmunoassay, and chromatographic procedures using a variety of region-specific antibodies recognizing separate portions of probombesin. In situ hybridization using cRNA probes was employed to analyze bombesin gene expression at a cellular level. A novel procedure using a divalent form of bombesin and gold-labeled monoclonal antibodies for the localization of bombesin binding sites at the ultrastructural level was employed in this study. Antibodies to neuron-specific enolase and electron microscopy were employed for the determination of neuroendocrine differentiation. Surgical samples of pulmonary (n = 250) and nonpulmonary (n = 28) small cell carcinomas, 49 carcinoids, and 62 atypical lung carcinoids were investigated and compared with 169 control tumors, including lymphomas, adenocarcinomas, squamous cell carcinomas, and non-small-cell undifferentiated tumors. Cell lines cultured from pulmonary small cell carcinoma and smear preparations of pleural effusions from patients with small cell carcinoma of the lung were also investigated. Strong immunostaining for neuron-specific enolase was noted in all neuroendocrine tumors investigated, and no immunoreactivity was noted in control cases. Electron-dense neurosecretory granules were abundant in carcinoid tumors, scattered in small cell carcinoids, and absent in control cases. Immunostaining for bombesin was particularly strong in benign carcinoids, whereas the more malignant neuroendocrine tumors (e.g., small cell carcinomas) stained best with antibodies to the carboxyl-terminal flanking portion of human probombesin (proGRP). These findings were further validated by radioimmunoassay and chromatography of tissue extracts. Specific binding sites for bombesin were demonstrated on the surface of small cell carcinoma cells maintained in culture. In situ hybridization demonstrated mRNA for preprobombesin in all small cell carcinomas investigated, including surgical samples, cytological preparations, and cell lines. Hybridization reactions varied in intensity, with some cells in autoradiograms almost masked by silver grains and others showing much lighter deposits.

Bombesin↗

PubMed1988

Expression of the atrial natriuretic peptide gene in the cardiac muscle of rat extrapulmonary and intrapulmonary veins.

Atrial natriuretic peptide is a peptide regulating salt and water balance, originally isolated from the cardiac atrium, where it is synthesised as part of a precursor molecule in specialised myocardial cells. The myocardium extends into the extrapulmonary part of the pulmonary veins in many species, including man. In some small mammals, however, such as the rat, mouse, and bat, it extends further to veins in the peripheral parts of the lung. Since this myocardial layer is continuous with that in the atrium, we have looked for the possible expression of the atrial natriuretic peptide gene in this tissue in rats. Strong immunoreactivity was seen for both the peptide and the N terminal sequence (cardiodilatin) of its precursor in extrapulmonary veins and in intrapulmonary veins extending into the lung as far as the second branching point, where it was localised in the dense cored granules by electron microscopy; in situ hybridisation showed atrial natriuretic peptide messenger RNA at identical sites. Chromatography and radioimmunoassay of extracts of extrapulmonary and intrapulmonary veins showed most of the atrial natriuretic peptide immunoreactivity to be in the uncleaved (precursor molecule) form. Thus the peptide is synthesised in veins both outside and inside the lung, and these extra-atrial sites may be an important additional source of circulating atrial natriuretic peptide.

Animals↗

PubMed1987

Extraadrenal paragangliomas. An immunocytochemical and ultrastructural report.

Although the majority of extraadrenal paragangliomas are nonfunctional, some of these tumors are associated with hormone production and clinical symptoms, notably hypertension. The authors have investigated 22 paragangliomas, five of which were diagnosed as clinically functional in a light microscopic immunocytochemical and electron microscopic study (nine cases). Histologically, all the paragangliomas exhibited similar features, with a "Zellballen" pattern of polygonal cells. All 22 cases were strongly immunoreactive to protein gene product 9.5 (PGP 9.5) antisera and moderately reactive to antineuron-specific enolase (NSE) sera. Ten cases (five functional) were focally immunoreactive to antichromogranin sera. Seven cases (four functional) were immunoreactive to neuropeptide Y and enkephalin antisera, and six (five functional) to tyrosine hydroxylase antisera. The clinically functional tumors expressed at least two of the antigens, enkephalin, neuropeptide Y, or tyrosine hydroxylase, whereas none of the 17 nonfunctional possessed more than one of these. Electron microscopic study revealed cells from all the nine cases studied to contain secretory granules. Granule sizes ranged from 100 to 280 nm and the morphologic examination of the secretory granules generally showed a dense core with a membrane-bound halo of variable size. Secretory granules were observed in the five functional cases and these were larger (220-280 nm) than those seen in the nonfunctional tumor cells (100-180 nm). Also, tumor cells from the functional cases contained numerous dilated mitochondrial profiles.

Adult↗

PubMed1987

Immunohistochemical markers of small cell carcinoma and related neuroendocrine tumours of the lung.

A selected group of 263 pulmonary neuroendocrine tumours comprised 156 small cell carcinomas, five combined cell carcinomas, nine atypical carcinoid/small cell carcinomas, 32 atypical carcinoids, ten large cell/small cell carcinomas, and 51 carcinoid tumours. These were compared with a group of 109 non-small cell carcinomas, using four markers of neuroendocrine differentiation to determine differences in reactivity between the two groups and among the variants of neuroendocrine tumour. The antibodies used were neuron-specific enolase (NSE), protein gene product (PGP) 9.5, human bombesin, and the C-terminal flanking peptide of human bombesin (CTP). Most small cell carcinomas, carcinoid tumours, and atypical carcinoid variants showed immunoreactivity for both NSE and PGP 9.5 but a significant number of non-small cell carcinomas, mainly squamous cell carcinomas, were also positive (11 and 35 per cent, respectively). Bombesin was specific for neuroendocrine tumours, being demonstrable in 35 per cent carcinoids and 24 per cent small cell carcinomas, but staining was focal and often confined to scattered cells. Diffuse strongly positive immunoreactivity for CTP was seen in the majority of malignant neuroendocrine tumours, but only 12 per cent of carcinoid tumours were positive and non-small cell carcinomas were negative. CTP is therefore of potential value as a specific marker of malignant neuroendocrine tumours, particularly if the amount of biopsy material is limited and the tumour is an unusual variant, such as atypical carcinoid or large cell-small cell carcinoma.

Biomarkers, Tumor↗

PubMed1987

Localization of atrial natriuretic peptide mRNA and immunoreactivity in the rat heart and human atrial appendage.

The localization of mRNA encoding preproatrial natriuretic peptide was investigated in tissue sections and cultures of rat heart and in sections of human right atrial appendage using the technique of in situ hybridization with 32P- and 35S-labeled RNA probes. Rat atrial natriuretic peptide (ANP) transcripts were demonstrated in numerous atrial myocytes and, to a lesser extent, in ventricular myocytes in both tissue sections and newborn rat heart cultures. These findings are consistent with those obtained by RNA blot analysis of rat heart total RNA, indicating that a single prepro-ANP transcript of approximately 900 nucleotides was present in the ventricles as well as the atria. Using a 35S-labeled RNA probe for human ANP mRNA, ANP transcripts were also localized to the majority of myocytes in the human right atrial appendage. Only background levels of autoradiographic labeling were obtained when RNA probes identical to the coding sequence of rat or human ANP mRNA were used. A close correlation was found between the distribution of ANP immunoreactivity and prepro-ANP mRNA in these preparations. These results provide unequivocal evidence for the expression of the ANP gene in the rat ventricles, as well as the atria, because myocytes in these tissues have been established as the sites of both ANP localization and precursor biosynthesis. The combined use of cardiac cultures and in situ hybridization may be of value in future studies investigating the regulation of ANP synthesis in cardiac myocytes.

Animals↗