BRIEF REPORT |
Correspondence to: Nicole Mahy, Unitat de Bioquímica, Facultat de Medicina, U.B., C/ Casanova 143, E-08036 Barcelona, Spain.
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Summary |
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We studied monoamine oxidase (MAO) A and B localization in human pancreas, thyroid gland, and adrenal gland by immunohistochemistry. The primary antibodies used were mouse monoclonal anti-human MAO-A (6G11/E1) and anti-human MAO-B (3F12/G10/2E3). Samples were obtained from six routine autopsy cases and fixed in 2% paraformaldehyde. Exocrine pancreas showed a widespread distribution of MAO-A, whereas MAO-B was present only in centroacinar cells and epithelial cells of pancreatic ducts. In endocrine pancreas, MAO-A was observed in around 50% of islet cells, whereas MAO-B was less abundant and was restricted to the periphery of islets. Thyroid gland showed strong MAO-A immunoreactivity in all cell types and was MAO-B-negative. In adrenal gland, the capsule displayed MAO-A but not MAO-B immunoreactivity, whereas the cortex showed widespread MAO-A staining but was MAO-B-negative in interstitial cells. Finally, in the medulla only a few scattered cells showed either MAO-A or MAO-B immunoreactivity. To our knowledge, these data represent the first study of the cellular distribution of MAO-A and MAO-B in the three human tissues included. (J Histochem Cytochem 48:147151, 2000)
Key Words: monoamine oxidases (MAOs), pancreas, thyroid gland, adrenal gland, human, immunohistochemistry
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Introduction |
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Monoamine oxidases (MAOs; EC 1.4.3.4) are the main degradative enzymes of monoamine hormones and neurotransmitters such as epinephrine, norepinephrine, serotonin (5-HT), and dopamine. Two MAO subtypes, named MAO-A and MAO-B, have been identified (
In an attempt to understand the role of MAO in the brain, many studies have investigated the localization of MAO-A and MAO-B in the human brain. In contrast, little is known about their distribution in peripheral tissues, especially in humans. In a recent paper we described the distribution of MAO-A and MAO-B in six human peripheral tissues by means of quantitative autoradiography (
Samples of pancreas, adrenal gland, and thyroid gland were generously provided by Dr. J.A. Bombí (Departament de Biologia Cel·lular i Anatomia Patològica, Facultat de Medicina, Universitat de Barcelona, Spain) and were obtained from six routine autopsy cases (three men and three women) and taken in accordance with European guidelines. Tissues affected by the pathological state of the patient were discarded. The age range was 5084 years (mean ± SEM 73.6 ± 4.9) and the postmortem delay ranged from 4 to 22 hr (mean ± SEM 12.3 ± 2.5).
Immediately after autopsy, tissues were fixed by immersion in 2% (w/v) paraformaldehyde in 0.1 M phosphate buffer (pH 7.4) at 4C for 4 hr. After fixation, tissues were immersed for 72 hr in 15% (w/v) sucrose in 0.1 M phosphate buffer for cryoprotection, frozen with dry ice, and stored at -30C.
Cryostat sections (12 µm) were obtained and mounted on gelatinized slides. The immunohistochemical peroxidaseanti-peroxidase (PAP) technique was used. Endogenous peroxidase activity was inhibited by a 30-min preincubation in H2O2methanolPBS (0.01 M phosphate buffered saline) (0.3/9.7/90) followed by a 10-min wash in PBS. As primary antibodies for detection of MAO-A and MAO-B protein, the mouse monoclonals anti-human MAO-A (6G11/E1) (see Figure 1) and anti-human MAO-B (3F12/G10/2E3) (-gliadin, which has been typed as subclass IgG1) was diluted 1:50 in normal swine serum and used to define the nonspecific staining. After washing, sections were incubated for 30 min in rabbit anti-mouse IgG (1:100), rinsed and washed in PBS, and incubated for 30 min in mouse IgGPAP (1:100). Sections were developed for 15 min in a 0.05 M Tris solution containing 0.03% diaminobenzidine and 0.006% H2O2. Some sections were counterstained with Mayer's hematoxylin. All antibodies except the primary ones were purchased from Sigma (St Louis, MO).
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In the exocrine pancreas, strong MAO-A immunoreactivity (ir) was seen in all acinar and centroacinar cells, in the intercalated ducts, and in the epithelial cells of pancreatic ducts. The walls of pancreatic ducts and the connective tissue showed no MAO-A ir (Figure 2A and Figure 2B). In contrast, acinar cells were devoid of MAO-B ir. Centroacinar cells showed punctate MAO-B staining, and both intercalated ducts and epithelial cells of pancreatic ducts were enriched in MAO-B ir. Connective tissue was devoid of MAO-B ir (Figure 2C).
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In the endocrine pancreas, both MAO-A and MAO-B ir was present but in different cell populations (Figure 2D2F). MAO-A-positive cells were homogeneously distributed in the islets. They accounted for approximately 50% of the islet cells and probably correspond to ß-cells. MAO-B-positive cells were less abundant, appeared in the periphery of the islets, and may correspond to -cells.
In the thyroid gland, all follicular and parafollicular cells showed strong MAO-A immunoreactivity but were MAO-B-negative (Figure 2G2I).
In the adrenal gland, the capsule showed MAO-A but not MAO-B ir (Figure 2J2L). Positive staining was seen for both isoenzymes in the secretory cells of the cortex, but only interstitial cells displayed MAO-A ir. The only MAO-A immunoreactivity in the medulla was observed in the walls of the central vein and in scattered cells. The pattern of MAO-B ir in the medulla was similar to that of MAO-A (Figure 2L).
In this article we report the cellular distribution of MAO-A and MAO-B in human pancreas, thyroid, and adrenal glands by immunohistochemistry. To our knowledge, this is the first study that characterizes the cellular distribution of MAO isoenzymes in these tissues.
Acinar cells and centroacinar/duct cells participate in the production of pancreatic juice from the exocrine pancreas. We found MAO-A in both cell populations but MAO-B only in the latter. Because the sympathetic innervation of the exocrine pancreas is mainly if not entirely vascular, sympathetic norepinephrine is unlikely to be a MAO substrate in these cells. On the basis of rat experimental data, circulating monoamines, especially epinephrine, dopamine and 5-HT are the most likely MAO substrates in these cells.
In the endocrine pancreas, we have shown a different cellular distribution for the two isoenzymes. MAO-A is localized mainly in ß-cells, whereas MAO-B appears to be restricted to -cells. Of the three tissues included in this study, this is the only example of a cell type expressing MAO-B but not MAO-A, a feature also shared by platelets.
Both circulating epinephrine and sympathetic norepinephrine stimulate glucagon secretion and inhibit insulin secretion, and both are putative substrates for islet MAOs. However the pattern of distribution of MAO-A does not match that of norepinephrine found in all islet cells (
5-HT is another possible substrate for islet MAO. Because of multiple species-related differences regarding 5-HT and the endocrine pancreas (e.g., -cells, the role of MAO-B may be to prevent the accumulation of other amines that might interfere with 5-HT, as suggested for MAO-B in serotonergic neurons.
There are two types of parenchymal cells in the thyroid gland: the follicular cell, which synthesizes and secretes thyroxin (T4) and triiodothyronine (T3), and the parafollicular cell, which synthesizes and secretes calcitonin. Previous studies have shown that MAO-A is the predominant MAO isoform in the human thyroid gland (more than 90% of total MAO activity) (
The role of MAO-A in follicular cells is unclear. It has been suggested that it may supply the H2O2 required in thyroid hormone synthesis (
The detection of MAO-A but not MAO-B in parafollicular cells is a new finding. It is consistent with the reported presence of MAO-A, but not MAO-B, in human medullary thyroid carcinoma (MTC) cells, a cell line derived from a carcinoma of parafollicular cells. 5-HT has also been proposed as an MAO-A substrate in parafollicular cells, in which it may stimulate calcitonin secretion. However, an inhibitory effect has been proposed for intracellular 5-HT (
The adrenal gland is the main producer of hormonal catecholamines. However, we found neither MAO-A nor MAO-B ir in adrenal medullary cells. Nevertheless, we found MAO-A ir in the central vein and ganglion cells, which agrees with the findings of
In the adrenal cortex we found a more widespread distribution, with both MAO-A and -B ir present in all secretory cells. This is also the case in rat tissue, in which both enzymes have specific functions. Thus, the autocrine effect of dopamine synthesized from circulating L-dopa could be regulated by MAO-B, whereas MAO-A may metabolize the neuronal norepinephrine that modulates cortical endocrine release (
In conclusion, this study offers the first comprehensive results showing that MAO-A and MAO-B are localized at the cellular level in these human peripheral tissues. They show a widespread localization of both enzymes, especially MAO-A, and frequent co-localization. These observations are in marked contrast to the pattern in brain, in which most cells are devoid of both isoenzymes (e.g., oligodendrocytes, microglia, and most neurons) and co-localization of MAO-A and MAO-B is extremely rare. The physiological role of the enzymes in pancreas, thyroid gland, and adrenal gland is far from being understood, but these results suggest that specific uptake or synthesis of monoamines occurs in cell types in which MAO is present. Further experiments are required to determine the physiological substrates for MAO-A and MAO-B in the different cell types.
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Acknowledgments |
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Supported by the Wellcome Trust and FISss 94/1461.
Received for publication June 3, 1999; accepted August 18, 1999.
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