The use of a screening test for thyroid functional disorder by sensitive thyroid stimulating hormone assay in the elderly was investigated. The basal thyroid stimulating hormone levels predicted the response of thyroid stimulating hormone to thyrotropin releasing hormone; it was suppressed in 99 (99.0%) of 100 hyperthyroid patients. Therefore, not only primary hypothyroidism but also hyperthyroidism can be excluded when the serum thyroid stimulating hormone levels are normal. An epidemiological study was then performed on 2,421 (76.7%) of the Japanese general population aged 40 or over recruited from the residents in Hisayama town and also in 122 residents between 20 and 40 years of age. Additional free T4 measurement was necessary in about 10% of the residents with abnormal TSH levels to confirm the diagnosis of hyperthyroidism or distinguish latent from overt hypothyroidism. There was a significant correlation between age and serum thyroid stimulating hormone levels after logarithmic conversion (r = 0.1533, P less than .001). The prevalence of thyroid dysfunction found in 1,026 males and in 1,395 females aged 40 or over was, respectively: hyperthyroidism, less than 0.1% and 0.2%, latent (subclinical) hypothyroidism, 3.2% and 5.5%, and overt hypothyroidism, 0.4% and 0.7%. We conclude that the screening with this sensitive thyroid stimulating hormone assay and additional free T4 measurement is useful for detection of patients with thyroid functional disorder.

The participation of lysosomal enzymes, hydroxyl radicals, and mitochondrial respiration in the cytocidal effect of TNF on tumor cells was investigated. The cytotoxicity of TNF on L-M cells was clearly reduced by lysosomotropic agents, DMSO (hydroxyl radical scavenger), NDGA (lipoxygenase inhibitor), and sodium azide (mitochondrial respiration inhibitor). The results suggest that lysosomal enzyme and hydroxyl radicals play an important triggering role in the destruction of tumor cells by TNF, and that the process of destruction might require ATP.

Several aspects of the activity and effects of tumor necrosis factor (TNF) were investigated to gain further insight into its cytotoxic mechanism. The relation between number of TNF receptors and TNF susceptibility of both tumor cells and normal cells was studied, utilizing a specific binding assay. Among the tumor cells, a fairly close correlation (r = 0.855) was observed between receptor number and sensitivity to TNF. No cytotoxic effect by TNF was observed on any of the normal cells tested, even though TNF receptors were shown to be present, and cell proliferation was apparently stimulated by TNF in some cases. TNF internalization and intracellular distribution were studied by pulse-labelling and Percoll density gradient centrifugation. In L-M (murine tumorigenic fibroblasts, highly sensitive to TNF cytotoxicity) cells and HEL (human embryonic lung cells, non-sensitive to TNF cytotoxicity) cells, receptor-bound 125I-labelled recombinant human TNF was rapidly internalized and delivered to lysosomes within 15-30 min, and this was followed by degradation and release into the culture medium. The presence of either a cytoskeletal disrupting agent or a lysosomotropic agent was observed to inhibit the cytotoxic effect of TNF, thus also indicating that TNF internalization, followed by delivery to lysosomes, is essential in the cytolytic mechanism of TNF. As observed by [3H]uridine incorporation, TNF did not affect RNA synthesis in L-R cells (TNF-resistant cell lines derived from L-M cells) and HEL cells, but markedly stimulated (by 3.5 times) RNA synthesis in L-M cells.

A synergistic increase in the cytotoxic effects of recombinant human tumor necrosis factor (rH-TNF) and anti-cancer drugs was demonstrated in vitro. The cytotoxicity of rH-TNF against L-M cells in combination with Mitomycin C (MMC), Adriamycin (ADM), Cytosine arabinoside (Ara-C), Actinomycin D (ACD), Daunomycin (DM), Cisplatin (CDDP), Vincristine (VCR), and 5-Fluorouracil (5FU), based on the concentration necessary for 50% inhibition of cell growth (IC50), was 4 to 347 times as high as that of rH-TNF alone. The results suggest that combination therapy including rH-TNF and anti-cancer drugs may be of value in the treatment of malignancy in human patients.

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