Lack of carcinogenicity of lyophilized Agaricus blazei Murill in a F344 rat two year bioassay.

The Brazilian mushroom Agaricus blazei Murill has antimutagenic, antioxidant, immunostimulatory and antitumorigenic activities, and is increasingly consumed as a health food worldwide. We undertook the present study to evaluate the chronic toxicity and oncogenicity of A. blazei Murill in F344 rats. To establish a no-observed-adverse-effect level (NOAEL), four treatment groups of 100 rats each (50 males and 50 females) were fed a powder diet containing lyophilized A. blazei aqueous extract at 0, 6250, 12,500, and 25,000 ppm for up to 2 years. During this period, there was no remarkable change in mean body weight, body weight gain, hematologic or serum chemistry parameters, or absolute or relative organ weights in control or treatment groups. Mortality in male treatment groups (26%, 16%, and 30%), however, was significantly lower than in controls (48%). Histopathological studies showed no increased incidence of tumors in any treatment group, and total tumor incidence across all groups was comparable to historical data. In conclusion, an A. blazei Murill lyophilized powder diet even at 25,000 ppm (1176 mg/kgb x w x /day for male rats and 1518 mg/kgb.w./day for female rats) resulted in no remarkable carcinogenic effects in F344 rats over a 2-year period. Therefore, the dietary NOAEL is 25,000 ppm.

Natural killer cell activity and quality of life were improved by consumption of a mushroom extract, Agaricus blazei Murill Kyowa, in gynecological cancer patients undergoing chemotherapy.

A mushroom extract, Agaricus blazei Murill Kyowa (ABMK), has been reported to possess antimutagenic and antitumor effects. Here, we investigate the beneficial effects of ABMK consumption on immunological status and qualities of life in cancer patients undergoing chemotherapy. One hundred cervical, ovarian, and endometrial cancer patients were treated either with carboplatin (300 mg / m(2)) plus VP16 (etoposide, 100 mg / m(2)) or with carboplatin (300 mg / m(2)) plus taxol (175 mg / m(2)) every 3 weeks for at least three cycles with or without oral consumption of ABMK. We observed that natural killer cell activity was significantly higher in ABMK-treated group (ANOVA, n = 39, P < 0.002) as compared with nontreated placebo group (n = 61). However, no significant difference in lymphokine-activated killer and monocyte activities was observed in a manner similar to the count of specific immune cell populations between ABMK-treated and nontreated groups. However, chemotherapy-associated side effects such as appetite, alopecia, emotional stability, and general weakness were all improved by ABMK treatment. Taken together, this suggests that ABMK treatment might be beneficial for gynecological cancer patients undergoing chemotherapy.

Protective effects of green tea extracts (polyphenon E and EGCG) on human cervical lesions.

We investigated clinical efficacy of green tea extracts (polyphenon E; poly E and (-)-epigallocatechin-3-gallate [EGCG]) delivered in a form of ointment or capsule in patients with human papilloma virus (HPV) infected cervical lesions. Fifty-one patients with cervical lesions (chronic cervicitis, mild dysplasia, moderate dysplasia and severe dysplasia) were divided into four groups, as compared with 39 untreated patients as a control. Poly E ointment was applied locally to 27 patients twice a week. For oral delivery, a 200 mg of poly E or EGCG capsule was taken orally every day for eight to 12 weeks. In the study, 20 out of 27 patients (74%) under poly E ointment therapy showed a response. Six out of eight patients under poly E ointment plus poly E capsule therapy (75%) showed a response, and three out of six patients (50%) under poly E capsule therapy showed a response. Six out of 10 patients (60%) under EGCG capsule therapy showed a response. Overall, a 69% response rate (35/51) was noted for treatment with green tea extracts, as compared with a 10% response rate (4/39) in untreated controls (P<0.05). Thus, the data collected here demonstrated that green tea extracts in a form of ointment and capsule are effective for treating cervical lesions, suggesting that green tea extracts can be a potential therapy regimen for patients with HPV infected cervical lesions.

Chemopreventive effect of green tea (Camellia sinensis) against cigarette smoke-induced mutations (SCE) in humans.

Green tea (Camellia sinensis) is consumed daily between the meals or after meals in Japan and other Asian countries. In recent years, green tea and its major polyphenolics have been demonstrated to prevent chemically induced tumors in a variety of experimental animal models system. The exact mechanism(s) of its anticarcinogenic activity remains to be elucidated, but green tea polyphenolics have demonstrated antimutagenic, anticarcinogenic, antioxidant, and antipromotional effects, including inhibition of Phase I and inducing Phase II enzymes. Enzyme activities of glutathione peroxidase, catalase, and quinone reductase, and glutathione S-transferase are also induced. However, a paucity of green tea effects in humans prompted us to investigate antimutagenic effects of green tea against smoke-induced mutation in humans. Chemopreventive effects of green tea and coffee among cigarette smokers were examined in 52 clinically healthy male subjects between 20-51 years of age. Blood specimens were obtained from non-smokers (Group I), smokers (II), smokers consuming green tea (III), and smoker/coffee drinkers (IV). The mean years of cigarette smoking (> 10 cigarettes/day) of Groups II, III, and IV ranged from 13.4-14.7 years. Daily intake of green tea and coffee was 3 cups/day/6 months (III and IV). The frequencies of sister-chromatid exchange (SCE) in mitogen-stimulated peripheral lymphocytes from each experimental group were determined and statistically analyzed. SCE rates were significantly elevated in smokers (9.46 +/- 0.46) vs. non-smokers (7.03 +/- 0.33); however, the frequency of SCE in smokers who consumed green tea (7.94 +/- 0.31) was comparable to that of non-smokers, implying that green tea can block the cigarette-induced increase in SCE frequency. Coffee, by contrast, did not exhibit a significant inhibitory effect on smoking-induced SCE.

Case report: rupture and growth of adrenal myelolipoma in two patients.

Two cases of pathologically confirmed adrenal myelolipoma are described in Chinese patients. The first patient presented with tumour rupture and perinephric haemorrhage. The second patient was operated on 18 months after the incidental discovery of the tumour, due to its large size and the interval growth.

Chemopreventive effect of green tea (Camellia sinensis) among cigarette smokers.

Chemopreventive effects of green tea and coffee among cigarette smokers were examined in 52 clinically healthy male subjects between 20 and 52 years of age. Blood specimens were obtained from nonsmokers (group I), smokers (group II), smokers consuming green tea (group III), and smokers drinking coffee (group IV). The mean number of cigarette smoking years (> 10 cigarettes/day) in groups II-IV ranged from 13.4 to 14.7 years. Daily intake of green tea and coffee was 2-3 cups/day for 6 months (groups III and IV). The frequencies of sisterchromatid exchange (SCE) in mitogen-stimulated peripheral lymphocytes from each experimental group were determined and analyzed statistically. SCE rates were elevated significantly in smokers (9.46 +/- 0.46) versus nonsmokers (7.03 +/- 0.33); however, the frequency of SCE in smokers who consumed green tea (7.94 +/- 0.31) was comparable to that of nonsmokers, implying that green tea can block the cigarette-induced increase in SCE frequency. Coffee, in contrast, did not exhibit a significant inhibitory effect on smoking-induced SCE.

Selected testicular enzymes as biochemical markers for procarbazine-induced testicular toxicity.

Single doses of procarbazine (MIH) were injected IP at 0, 50, 100, 200, and 400 mg/kg body weight to CD-1 male mice. Activities of hyaluronidase, lactate dehydrogenase isoenzyme-X, and the dehydrogenases of sorbitol, alpha-glycerophosphate, glucose-6-phosphate, malate, isocitrate, and glyceraldehyde-3-phosphate in the testes of the mice were determined and correlated with changes in spermatogenic cell types in seminiferous tubules. All enzyme activities were higher than controls or remained unchanged on days 10-20 after drug treatment. Activities of hyaluronidase, sorbitol dehydrogenase, and lactate dehydrogenase isoenzyme-X decreased significantly to below normal levels on day 30 after drug treatment for all doses, whereas those of the other five dehydrogenases remained significantly higher than controls. All enzyme activities approached control levels with the concomitant recovery of spermatogenesis by day 60 after drug treatment. Histological examination of seminiferous tubules revealed that premeiotic spermatocytes were significantly reduced on days 10-20 but reappeared on day 30 after MIH treatment (400 mg/kg). The postmeiotic spermatogenic cells were unaffected at the time of MIH treatment, but had disappeared completely on day 30 after drug treatment. MIH, at the highest dosage, selectively destroyed spermatogonia and premeiotic spermatocytes; however spermatozoa and elongated spermatides were unaffected. This study demonstrated that the cytotoxic effect of MIH on spermatogenesis could be evaluated via changes in testicular enzyme activities. The present studies demonstrated that hyaluronidase, sorbitol dehydrogenase, and lactate dehydrogenase isoenzyme-X could serve as useful biochemical markers for assessing testicular toxicity induced by drugs and chemicals.

Aspects of testicular toxicity induced by anticancer drugs.

Newer histopathologic techniques were used in combination with sperm head counts (SHC) and serial mating (SM) studies to assess different aspects of testicular toxicity. Adult male rats were treated once intraperitoneally (i.p.) with vincristine (Vin: 0.15 and 0.6 mg/kg) or procarbazine (Proc: 50 and 200 mg/kg). Investigations were performed at weekly intervals until 10 weeks after treatment. SHC are a good parameter for cytotoxicity (Vin, Proc) assessed in great detail by morphological examination. Optimal fixation after perfusion and high optical resolution of semi-thin sections also allow the study of early and subtle specific alterations. However, they do not replace SM studies for the assessment of genotoxicity (Proc). In turn, SM studies are poor indicators of cytotoxicity.

Development of the acrosome and alignment, elongation and entrenchment of spermatids in procarbazine-treated rats.

Intraperitoneally administered procarbazine caused, among other features previously reported (Russell et al., 1983), specific defects in the acrosome of cap phase spermatids of the rat seminiferous epithelium. The effect of procarbazine was to fragment and eventually cause resorption of the acrosomes of a small number of steps 5--9 spermatids. Although the acrosome was lost, close union of the leaflets of the nuclear envelope underlying the acrosomal sac was maintained as was the marginal fossa and acrosomal zonule. Spermatids at steps 8 and 9 of development, which had lost their acrosomes, showed nuclei which were eccentric within the cell--a feature which normally occurs at these steps of spermiogenesis in acrosome intact cells. Even without an acrosomal sac, the plasma membrane of these cells (in stage VIII) became orientated to the region of the nuclear membrane which would have underlaid the acrosome. Although abundant, Sertoli ectoplasmic specialization did not become aligned with the spermatid head. The spermatid failed to become orientated within the seminiferous epithelium and failed to enter the crypts within the Sertoli cell as usually occurs during the elongation process. Thus, the presence of an acrosome is not likely related to the formation of an eccentric nucleus or the alignment of the surface of the nucleus which would normally underlay the acrosome with the cell's plasma membrane (internal alignment). The presence of an acrosome may be related to the alignment of the spermatid head with the ectoplasmic specialization, which in turn may influence the orientation and positioning of the late spermatids within the seminiferous epithelium (external alignment) and their position within recesses of the Sertoli cell. This study also suggests a role for the manchette in the process of elongation of the spermatid.

Morphological pattern of response after administration of procarbazine: alteration of specific cell associations during the cycle of the seminiferous epithelium of the rat.

Procarbazine, an anti-cancer agent, administered intraperitoneally to adult, male rats induced a characteristic morphological pattern of response in the seminiferous epithelium. Seminiferous tubules of rats receiving 100 mg/kg procarbazine and higher dosages displayed spermatids which were less mature than those normally found within seminiferous tubules which show a particular cell association. Early spermatids in steps 1-7 of spermiogenesis appeared arrested in their development and were present in cell associations which had advanced normally. The most probable cause of this apparent germ cell arrest was a retardation of acrosomal development since procarbazine is known to affect RNA and consequently protein synthesis. Other features which indicated defective RNA synthesis were the presence of abnormal spermatid nucleoli and abnormally configured chromatoid bodies. This study demonstrates, in contrast to what is indicated by present dogma, that apparent and temporary germ cell arrest may occur under certain deleterious conditions. It also illustrates that particular cell types within a cell association may be out of synchrony with the remainder of the cells in a cell association.

Studies on the male reproductive toxicity of Freon 22.

Freons have been used extensively as refrigerants and as propellants in household products, and yet their possible effects on male reproduction have received little attention. In the present study, adult male Sprague-Dawley rats (nine weeks of age) were exposed to 50 000 ppm Freon 22, five hrs per day for eight weeks. The control group received filtered air at an identical flow rate. At the end of the eight week exposure period, body and organ weights, hematology, blood chemistry, plasma gonadotropins, and fertility parameters were not significantly different from controls, with the exception of serum cholesterol levels, which were slightly higher, and glucose and triglyceride levels which were lower. The weight of coagulating glands was also lower than those of controls, but did not interfere with fertility function.

Differential DNA-repair activity in prespermiogenic cells of various mouse strains.

The present report demonstrates differential DNA-repair activity among 14 strains of immature (20 +/- 2 days old) male mice (inbred strains: C57BL/6J, RF/J, Nude homo/nu, RIII/2J, PL/J, AKR/J, Nude hetero/nude, C3H/HeJ, SWR/J, SM/J, ST/J, LP/J, BALB/cJ and random-bred strain: CD-1). The prespermiogenic cells were isolated and enriched by collagenase-trypsin digestion of seminiferous tubules and subsequent 3% albumin-gradient centrifugation. Enriched prespermiogenic cells demonstrated a viability greater than 95% by trypan blue exclusion criteria. For in vitro unscheduled DNA synthesis (UDS) determination, prespermiogenic cells (10(6) cells/ml) were incubated with methyl methanesulfonate (0.4 mM) in the presence of 20 mM hydroxyurea (HU). At 20 mM HU concentration, 90% of S-phase DNA activity in prespermiogenic cells was inhibited and thus, the net UDS activity following MMS exposure was readily determined. MMS-induced UDS activity in the CD-1 mouse strain was both linear up to 4 h of incubation and dose-dependent at 4 h of incubation. The apparent Km for MMS-induced UDS activity in prespermiogenic cells was approx. 1.8 x 10(-4) M. Of the 14 mice strains tested, C57BL/6J and RF/J exhibited the highest DNA-repair activity, while BALB/cJ, LP/J, and ST/J showed the lowest. A maximal difference in UDS activity of 3.5-fold was observed between C57BL/6J and BALB/cJ. Furthermore, a 2.5-fold difference was also noted between RF/J and LP/J mouse strains. Thus, wide variations in DNA-repair activity among 14 mouse strains were clearly demonstrated. Whether genetically select mouse strains with the lowest DNA-repair activity should have greater sensitivity toward environmental mutagens needs to be tested.

Induction of aryl hydrocarbon hydroxylase activity in the rat prostate glands by 2,3,7,8-tetrachlorodibenzo-p-dioxin.

Because of the interest in male reproductive tract toxicity, we determined the effects of 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) on cytochrome P-450 levels and the specific activity of aryl hydrocarbon hydroxylase (AHH), epoxide hydrase and glutathione S-transferase in the rat prostate glands after a single oral dose of TCDD (10 microgram/kg b.wt.). The maximum induction of AHH activity and cytochrome P-450 levels in the prostate glands was approximately 200- and 6.5-fold that of controls. After TCDD treatment, the peak induction of AHH activity and cytochrome P-450 levels in the prostate glands was observed at 16 and 48 hr, respectively; however, the specific activity of epoxide hydrase or glutathione S-transferase in prostate glands was not induced at any time after TCDD treatment. TCDD-induction of AHH and cytochrome P-450 contents can be completely inhibited by a single pretreatment with either actinomycin D or cycloheximide, suggesting TCDD induces de novo synthesis of specific proteins. Dramatic increases in AHH activity (200-fold increase) were associated with the new formation of cytochrome P-450 in the rat prostate glands after a single oral dose of TCDD. The biological significance of such a dramatic increase of prostatic AHH activity and the emergence of new cytochrome P-446 in rat prostate glands is unknown at the present time.