Effect of Ubiquinol on Serum Reproductive Hormones of Amenorrhic Patients.

In neuroendocrine system the increase in oxidative status is produced by a glucocorticoid-dependent and transcriptional increase in pro-oxidative drive, with concurrent inhibition of the antioxidant defense system, ultimately leading to increased neuronal cell death. Functional hypothalamic disturbances and neuroendocirne aberrations have both short and long term consequences for reproductive health. Understandably, an impaired or diminished hypothalamic-pituitary-ovarian axis leads to anovulation and hypoestrogenism. Anovulation is directly linked to the neurohormonal and hormonal background of Functional Hypothalamic Amenorrhea. Impairment of pulsatile Gonadotropin Releasing Hormone secretion causes the impairment of pulsatile Lutenizing Hormone (LH) and Follicle Stimulating Hormone (FSH) secretion. The importance of oxidative stress in various pituitary disorders suggesting a possible clinical usefulness of antioxidant molecules like the lipophilic antioxidant Ubiquinol. Coenzyme Q10 or Ubiquinol is an essential part of the cell energy-producing system of mitochondria. However, it is also a powerful lipophilic antioxidant, protecting lipoproteins and cell membranes from autooxidation. Due to these unique actions Ubiquinol is used in clinical practice as an antioxidants for neurodegenerative diseases. So to identify the role of Ubiquinol on reproductive hormones FSH and LH, we have included 50 infertile patients of age group of 20-40, which are mostly amenorrhic. Out of 50 only 30 patients were in continuous follow up after supplementing them with 150 mg of Ubiquinol every day for 4 months. The hormonal levels were estimated by Enzyme Linked Immuno Sorbent Assay technique at follicular phase. The result suggests that FSH concentration is increased up to three times (from 3.10 ± 2.70 to 10.09 ± 6.93) but remains within the normal limit (P < 0.05). LH values were found doubled (P < 0.05) than its normal range (from 14.83 ± 10.48 to 27.85 ± 22.30). The Prolactin values were decreased while Progesterone values were high but not in the significant range (P > 0.05). The supplementation of 150 mg of Ubiquinol may reduce the oxidative stress in neuroendocrine system which further improves the function of diminished HPA axis. Hence increased level of FSH and LH may be due to reduced oxidative stress by Ubiquinol.

Tumour growth inhibition in mice by glycosylated recombinant human lymphotoxin: analysis of tumour-regional mononuclear cells involved with its action.

We compared the antitumour effects of glycosylated LT (gLT), nonglycosylated LT and TNF against a solid tumour in mice. We found that: (a) The systemic administration of gLT showed significant antitumour activity. These effects were, however, quite small in nude mice. Nonglycosylated LT and TNF attained the same degree of effectiveness as gLT, but at a 5-times higher dose. The serum half-life of gLT was 3-fold longer than that of nonglycosylated LT and 22-fold longer than that of TNF. (b) The effect of gLT was significantly blocked by pretreatment with anti-asialo GM1 antibody. Treatment with gLT produced a significant reduction in numbers of tumour-regional mononuclear cells, which in turn, produced increases intensive necrosis. (c) Mononuclear cells in the tumour tissues before gLT-injection were predominantly IL-2 receptor +/CD3- cells and CD3+ cells. Pretreatment with the anti-asialo GM1 antibody produced a drastic reduction of IL-2 receptor +/CD3- cells. These findings suggest that the efficient antitumour effect of gLT is due to a longer serum half-life than that of nonglycosylated LT or TNF in vivo, and its function is largely mediated by IL-2 receptor +/CD3- cells.

Prevention of autoimmune diabetes with lymphotoxin in NOD mice.

We have reported previously that chronic and systemic administration of a streptococcal preparation (OK-432), an inducer of TNF, or of recombinant hTNF prevented the development of IDDM in the two animal models of IDDM-NOD mice and BB rats. In this study, we examined the effect of LT, which is structurally and functionally related to TNF, on NOD mice with diabetes. The cumulative incidence of diabetes at 30 wk of age was 22 of 40 (55%) in nontreated female NOD mice and was 4 of 8 (50%; NS), 3 of 29 (10%; P < 0.001), and 0 of 8 (0%; P < 0.001) in female mice treated three times a week from 4 to 30 wk of age with 5, 50, or 500 U of recombinant hLT, respectively. Intensity of insulitis was slightly reduced in the long-term LT-treated mice. LT productivity by ConA-stimulated spleen cells was examined in vitro. Although no significant difference was found between NOD mice and the other mouse strains, female NOD mice were slightly but significantly (P < 0.01) lower producers of LT immunoreactivity than male NOD mice, the diabetes incidence of which is lower than that of females. The SMLR as a marker of normal immune response, which was reported to be impaired in autoimmune animals including NOD mice, was significantly lower in female than male NOD mice. However, the low SMLR in female NOD mice was significantly increased by the administration of LT, and the increase was mediated by the responder cells of the LT-treated mice.(ABSTRACT TRUNCATED AT 250 WORDS)

Usefulness of glycosylated recombinant human lymphotoxin for growth inhibition of human and murine solid tumors and experimental metastasis in mice.

We have examined the antitumor and antimetastatic effects of native-type, glycosylated recombinant lymphotoxin (LT) on human and murine tumors transplanted in mice. The results reported here are as follows: (a) The in vivo antitumor spectrum of LT is not coincident with the in vitro study, and it has a wide antitumor spectrum and substantially inhibits the growth of human solid tumors, (b) When both syngeneic and nude mice are transplanted with Meth A tumor, the significant growth-inhibitory effect of LT is obtained in syngeneic mice, but the effect is quite small in nude mice regardless of the routes; LT attains the same degree of effectiveness as that in syngeneic mice, but at an 8 to 16 times higher dose. Furthermore, the pretreatment with anti-asialo-GM1 antibody inhibits the antitumor effects of LT in syngeneic mice, (c) In the pulmonary metastasis model induced by i.v. injection of Meth A cells, a high preventive effect of LT is obtained by systemic administration in syngeneic mice, but not in nude mice. In addition, the pretreatment with anti-asialo-GM1 antibody completely prevents the antimetastatic effect of LT, but also blocks that effect of control mice without LT treatment. In conclusion, LT appears to be a potent cytokine against tumor growth and metastasis in vivo. The differences between nude and syngeneic mice suggest the involvement of host immunity in the expression of LT function.

Synergistic antitumor effect of glycosylated recombinant human lymphotoxin with human interferon-gamma on lymphotoxin-sensitive human tumor.

We examined the antitumor effect of glycosylated recombinant lymphotoxin (LT) in combination with human interferon-gamma (IFN-gamma) on human tumors transplanted into nude mice and compared it with that of tumor necrosis factor (TNF). The results were as follows: (i) The systemic administration of glycosylated LT combined with IFN-gamma produced a significant antitumor activity against HT-1080 fibrosarcoma, G-361 malignant melanoma, KB nasopharyngeal carcinoma, and ZR-75-1 breast carcinoma, all of which are relatively resistant to a single treatment with LT or IFN-gamma. The synergistic effect was also seen in LT-sensitive HeLa S3 tumors. The effect was observed after either i.v. or s.c. injection. (ii) In contrast, no synergistic or additive effect on HeLa S3 tumors was observed in the case of TNF combined with IFN-gamma. (iii) The serum half-life of glycosylated LT in tumor-bearing mice was about 22-fold longer than that of TNF. In conclusion, glycosylated LT, especially in combination with IFN-gamma, appears to be a potent cytokine against tumor growth in vivo compared with TNF. Its long serum half-life can result in a strong antitumor effect in combination with IFN-gamma in vivo.

The pharmacokinetic pattern of glycosylated human recombinant lymphotoxin (LT) in rats after intravenous administration.

We have examined the pharmacokinetics of glycosylated recombinant human lymphotoxin (LT) after intravenous bolus injection in rats and compared them with those of tumor necrosis factor (TNF) or LT species. The results are as follows. 1) The mean half-life of glycosylated LT in serum increases for each increase in dose, and the distribution volume (V) and total body clearance [(Cl (total)] tend to decrease for increase in dose. On the other hand, the half-life of TNF also increases for increase in dose, but the V tends to increase for increase in dose and Cl (total) does not change. 2) The glycosylated LT distributes to all organs so far tested except brain, and tends to accumulate to kidney more than other tissues at 6 h after the injection. 3) Nonglycosylated LT produced by E. coli and the glycosylated LT species carrying both N-type and mucin-type sugar moieties (25 kDa LT) have shorter half-lives and higher Cl (total)s than 23 kDa LT carrying N-type sugar moieties alone. The 21 kDa LT, the same species as 23 kDa LT except that it lacks 15 amino acid residues at the N-terminus, disappears much faster than 23 kDa LT and shows higher V and Cl (total). Thus, glycosylated LT shows nonlinear pharmacokinetics like TNF, but the deposition is quite different from that of TNF. The high serum concentration of glycosylated LT depends upon the presence of N-type sugar moieties, but not mucin-type sugar moieties. The N-terminal protein chain of LT also correlates with the serum concentration.

Reversible induction of anchorage independent growth from normal mouse epidermal keratinocytes, MSK-C3H-NU, in soft agar medium by 12-O-tetradecanoylphorbol-13-acetate and epidermal growth factor.

The induction of anchorage independent growth occurred in nontumorigenic, mouse epidermal keratinocyte, MSK-C3H-NU cells when inoculated in a 0.3% soft agar medium with 12-O-tetradecanoylphorbol-13-acetate and/or epidermal growth factor. Colonies appeared about 3 weeks after incubation and keratinization of various forms occurred in them. 10-43 cells, a subline of higher subcultivation level, showed a rather sensitive response to these chemicals. 12-O-tetradecanoylphorbol-13-acetate and epidermal growth factor each induced anchorage independent growth but additive effects were also observed. Cell lines were established from clonal growths recovered from these induced colonies in soft agar. Morphologically, they retained their original normal clonal growth and keratinization patterns. Anchorage independent growth was not retained. Response to 12-O-tetradecanoylphorbol-13-acetate and epidermal growth factor, however, became more sensitive in some of the recovered cell lines when compared to their original cells. These data show that the induction of anchorage independent growth in soft agar is reversible in the MSK-C3H-NU cell system.