Faculty Bibliography

Reactive amyloidosis is a disease occurring in patients suffering from chronic infections, inflammation, and certain malignant conditions that are characterized by a considerable elevation of the acute phase reactant serum amyloid A (SAA). It is defined by the presence of extracellular deposits of fibrillar material containing amyloid A (AA) as its main component. AA is an 8.5-kd protein structurally identical to the NH2-terminal of the acute phase reactant SAA. SAA consists of a group of evolutionally conserved amphipathic proteins, encoded by a large number of genes and produced abundantly during inflammation, all suggesting an important role, probably of a neutralizing (anti-inflammatory) nature. An analysis of various aspects of SAA provides no clues to the mechanism of amyloid production, its occurrence in only selected individuals, and its preferential relationship to one isotype of SAA. Until more data is available, the present view on AA amyloidogenesis remains hypothetical.

The clinical course and pathological findings of five cases of amyloid goiter in patients with AA amyloidosis are presented. The underlying diseases which caused the amyloidosis were: chronic recurrent multifocal sterile osteomyelitis, acne conglobata, tuberculosis, and two cases of familial Mediterranean fever. All presented with a hard, moderately rapidly growing thyroid mass, mimicking a malignant thyroid tumor. Four patients underwent excisiorial biopsy or partial thyroidectomy, after which they developed hypothyroidism. The fifth case was discovered at an autopsy. Histologically, in all cases, the thyroid follicles were scarce, many had an atrophic appearance, surrounded by amyloid deposits and associated with adipose metaplasia of the stroma. The amyloid was potassium permanganate Congo red sensitive, and reacted with anti-AA antibodies by the avidin biotin immunoperoxidase method in all cases. The amyloid fibrils, extracted from the thyroid tissue of each of the five cases, revealed two protein bands in 17% SDS polyacrylamide gel electrophoresis: One band (of about 5 kD) ran faster than the usual 8.5 kD amyloid A protein, and the other (of about 9.5 kD) ran slower. Both proteins reacted with rabbit anti-human AA polyclonal antiserum in Western immunoelectroblot. It is concluded that in our cases amyloid goiter was associated with two SAA related protein chains, that were codeposited in the thyroid in the form of fibrils, accompanied by large amounts of mature adipose tissue. © 1995 Informa UK Ltd All rights reserved: reproduction in whole or part not permitted.

The determination of the amount of amyloid in tissues is important to follow the onset and the development of amyloidosis. In the present study a new procedure for quantification of amyloid proteins in AA amyloidosis is described. It involves the protein extraction from mouse and human tissue and evaluation of their amount by inhibition ELISA. The procedure is simple and may be performed using milligram amounts of tissue. The sensitivity limit of protein AA detection corresponds to 0.007mg/mg tissue. The results obtained using the inhibition ELISA were consistent with those obtained by Congo red staining. In contrast to previously developed amyloid quantification techniques, the presented procedure required only the basic equipment avail-able in most laboratories. © 1995 Informa UK Ltd All rights reserved: reproduction in whole or part not permitted.

To study the mechanism of amyloid deposition, the nature of amyloid proteins formed in experimental murine amyloidosis, was examined. Spleen specimens, 15-60 mg, were homogenized and extracted using aqueous acidic acetonitrile, in a recently developed procedure, making it possible to obtain amyloid proteins from minute amounts of tissue. The extracted material, 1.5-4 mg, was analysed by Western blotting and ELISA using antibodies recognizing differentially proteins AA and SAA. Two immunoreactive proteins of 8 and 12 KDa were isolated and subjected to amino acid analysis and N-terminal sequence determination. The results of immunochemical and chemical examination showed that the 8 and 12 KDa proteins represented proteins AA and SAA, respectively. The data obtained provide new direct evidence for SAA in tissues during murine amyloidogenesis.

To characterize the expression of the IGF-I system in the spleen and its role in spleen growth, we have studied the effect of hypophysectomy and the action of either GH or IGF-I treatment on the expression of several components of the IGF system in the rat. Female Sprague-Dawley rats were hypophysectomized (Hx) on postnatal day 50, and five animals each received twice-daily sc injections of saline, bovine GH (bGH; 84 micrograms/animal/day), or recombinant human IGF-I (rhIGF-I; 125 micrograms/animal/day) for 11 days. Compared to sham-operated controls, Hx animals exhibited a reduction in both body (192.6 +/- 5.6 g (mean +/- S.E.M.) vs. 268.6 +/- 6.0 g; P < 0.001) and spleen weights (0.42 +/- 0.03 g vs. 0.84 +/- 0.06 g; P < 0.001). The reduction in body and spleen weights in Hx animals was partially prevented by both bGH and rhIGF-I. Body weights were 234.2 +/- 5.3 g (P < 0.001) after bGH and 213.8 +/- 6.3 g (P < 0.05) after rhIGF-I. Spleen weights were 0.56 +/- 0.048 after bGH P < 0.01 and 0.53 +/- 0.05 g after rhIGF-I (P < 0.05). Serum GH and IGF-I levels were markedly reduced in Hx animals and bGH partially maintained IGF-I levels. Hypophysectomy reduced spleen IGF-I mRNA levels (30.6 +/- 7.5% of control values; P < 0.05) and this reduction was prevented by bGH (96.6 +/- 24.2%; NS) but not by rhIGF-I (39.9 +/- 5.0% NS vs. Hx). There were no changes in GH receptor or IGF-I receptor mRNA levels in Hx or bGH or rhIGF-I-treated animals. When IGF-I binding protein (IGFBP) mRNA levels were studied under these conditions, we found that IGFBP-1 mRNA was not detected in spleen; IGFBP-2 mRNA levels were reduced in Hx rats (67.9 +/- 7.4% of control values, P < 0.05) and bGH treatment prevented this reduction (95.5 +/- 12.2%, NS). IGFBP-3 mRNA levels were not affected by hypophysectomy or by bGH treatment, but were reduced in rhIGF-treated rats (69.6 +/- 3.0%, P < 0.05). On the other hand, IGFBP-4 mRNA levels were increased in Hx rats (136.4 +/- 15.9% of control values, P < 0.05) and bGH treatment prevented this increase.(ABSTRACT TRUNCATED AT 250 WORDS)

Growth and development of the spleen involves the growth hormone (GH)/insulin-like growth factor-I (IGF-I) axis. To evaluate the molecular mechanism of these effects we studied the effect of hypophysectomy (Hx) and GH replacement therapy on the expression of IGF-I, the IGF-I receptor and IGF-binding protein-2 (IGFBP-2) in juvenile rats. Hx resulted in a 30% reduction in body weight. GH replacement therapy for seven days partially prevented these effects. IGF-I mRNA levels were reduced 30% by Hx, IGFBP-2 mRNA levels fell 50% whereas IGF-I receptor mRNA levels were unaffected. GH therapy prevented the reduction in IGF-I and IGFBP-2 mRNA levels. These results suggest that the GH effect on splenic growth and development is via local (paracrine) IGF-I expression, in addition to any effect by circulating (endocrine) IGF-I.