Effect of Gosha-Jinki-Gan on Levels of Specific mRNA Transcripts in Mouse Testes after Busulfan Treatment.

With the increase in survival rates of cancer patients in recent years, infertility caused by anticancer treatments has become a significant concern for cancer survivors. Some studies have suggested that Sertoli cells play a key role in mediating testicular immunology in busulfan-induced aspermatogenesis. We recently demonstrated that Gosha-jinki-gan (TJ107), a traditional Japanese medicine, can completely recover injured spermatogenesis in mice 60 days after busulfan injection. In the present study, we sought to examine the levels of mRNA transcripts encoding markers of 25 Sertoli cell-specific products and 10 markers of germ cell differentiation. Our results demonstrated that only supplementation of TJ107 at day 60 after busulfan injection could significantly recover the increase in five mRNA species (Amh, Clu, Shbg, Testin, and Il1a) and the decrease in four mRNA species (Aqp8, CST9, Wnt5a, and Tjp1) in response to Busulfan (BSF) at day 120, with the increase of all examined spermatogenic markers.

Co-Administration of the Traditional Medicines Hachimi-Jio-Gan and Hochu-Ekki-To Can Reverse Busulfan-Induced Aspermatogenesis.

Busulfan is used as a chemotherapeutic drug to treat childhood and adult chronic myelogenous leukemia, and as an immunosuppressive agent before bone marrow transplantation. A key side effect of busulfan is the alteration of male reproductive function. Infertility caused by anti-cancer treatments has become a significant concern, but there are currently limited treatments for this condition. Recently, we demonstrated that Gosha-jinki-gan, a traditional Japanese medicine, completely reversed the spermatogenesis defects caused by cancer treatment in mice. Hochu-ekki-to and Hachimi-jio-gan are commonly used to treat male infertility, and Hachimi-jio-gan shares herbal ingredients with Gosha-jinki-gan. Therefore, in the present study, we administered Hachimi-jio-gan and Hochu-ekki-to alone or in combination to mice with severe aspermatogenesis caused by busulfan treatment. We performed testis weight measurements, quantitative histological assessments of the testes and the epididymis, and evaluated sperm counts and morphology. We also assessed the expression of immune mediators and macrophage markers. Treatment with a combination of both the medicines significantly reduced busulfan-induced testicular toxicity when compared to the lone treatment with either medicine. We demonstrated that treatment efficacy was related to a differential impact on testicular inflammation, and that the synergistic effect of co-administration completely reversed the busulfan-induced damage to the reproductive functions.

The effectiveness of traditional Japanese medicine Goshajinkigan in irradiation-induced aspermatogenesis in mice.

BACKGROUND: Infertility and gonadal dysfunction are well known side-effects by cancer treatment in males. In particularly, chemotherapy and radiotherapy induced testicular damage, resulting in prolonged azoospermia. However, information regarding therapeutics to treat spermatogenesis disturbance after cancer treatment is scarce. Recently, we demonstrated that Goshajinkigan, a traditional Japanese medicine, can completely rescue severe busulfan-induced aspermatogenesis in mice. In this study, we aimed to detect the effects of Goshajinkigan on aspermatogenesis after irradiation. METHODS: This is animal research about the effects of traditional Japanese medicine on infertility after cancer treatment. C57BL/6 J male mice received total body irradiation (TBI: a single dose of 6Gy) at 4 weeks of age and after 60 days were reared a Goshajinkigan (TJ107)-containing or TJ107-free control diet from day 60 to day 120. Then, two untreated females were mated with a single male from each experimental group. On day 60, 120 and 150, respectively, the sets of testes and epididymis of the mice in each group after deep anesthetization were removed for histological and cytological examinations. RESULTS: Histological and histopathological data showed that 6Gy TBI treatment decreased the fertility rate (4/10) in the control diet group; in contrast, in the TJ107-diet group, the fertility rate was 10/10 (p < 0.05 vs. 6Gy group). Supplementation with TJ107 was found to rescue the disrupted inter-Sertoli tight junctions via the normalization of claudin11, occludin, and ZO-1 expression and reduce serum anti-germ cell autoantibodies. CONCLUSIONS: These findings show the therapeutic effect on TBI-induced aspermatogenesis and the recovering disrupted gonadal functions by supplementation with TJ107.

Gosha-Jinki-Gan Recovers Spermatogenesis in Mice with Busulfan-Induced Aspermatogenesis.

Busulfan is an anti-cancer chemotherapeutic drug and is often used as conditioning regimens prior to bone marrow transplant for treatment of chronic myelogenous leukemia. Male infertility, including spermatogenesis disturbance, is known to be one of the side effects of anticancer drugs. While hormone preparations and vitamin preparations are used for spermatogenesis disturbance, their therapeutic effects are low. Some traditional herbal medicines have been administered to improve spermatogenesis. In the present study, we administered Gosha-jinki-gan (TJ107; Tsumura Co., Ltd., Tokyo, Japan) to mice suffering from severe aspermatogenesis after busulfan treatment to determine whether TJ107 can recover spermatogenesis. Male 4-week-old C57BL/6J mice were administered a single intraperitoneal injection of busulfan, and they were then fed a normal diet for 60 days and then a TJ107 diet or TJ107-free normal diet for another 60 days. After busulfan treatment, the weight of the testes and the epididymal sperm count progressively decreased in the normal diet group. On the other hand, in the TJ107 group, these variables dramatically recovered at 120 days. These results suggest that busulfan-induced aspermatogenesis is irreversible if appropriate treatment is not administered. Supplementation of TJ107 can completely recover the injured seminiferous epithelium via normalization of the macrophage migration and reduction of the expressions of Tool-like receptor (TLR) 2 and TLR4, suggesting that TJ107 has a therapeutic effect on busulfan-induced aspermatogenesis.

Preservation of rat limbs by hyperbaric carbon monoxide and oxygen.

Cold ischemia times ranging from <6 h to as long as 24 h are generally quoted as the limits for attempting the replantation of amputated extremities. In this study, we aimed to assess the effect of hyperbaric carbon monoxide (CO) and oxygen (O2) on rat limb preservation. Donor rat limbs were preserved in a chamber filled with hyperbaric CO and O2 for 3 days (CO + O2 3 days) or 7 days (CO + O2 7 days). Positive and negative control groups were created by using non-preserved limbs (NP) and limbs wrapped in saline-moistened gauze for 3 days (SMG 3 days), respectively. The survival rate of transplanted limbs at postoperative day 90 was 88% in the NP and 86% in the CO + O2 3 days. The corresponding survival rate was 50% in the CO + O2 7 days at postoperative day 90 but was 0% in the SMG 3 days at postoperative day 3. Muscle mass decreased in the CO + O2 3 days and CO + O2 7 days compared with the NP, but sciatic-tibial nerve conduction velocities did not differ. These results indicate that amputated extremities preservation with hyperbaric CO and O2 could extend the time limits of preservation, maintaining their viability for replantation.

Impulsive force on the head during performance of typical ukemi techniques following different judo throws.

In this study, eight judo athletes who are major candidates for the Japan national team were recruited as participants. Kinematic analysis of exemplary ukemi techniques was carried out using two throws, o-soto-gari, a throw linked to frequent injury, and o-uchi-gari. The aim of this study was to kinematically quantify the timing patterns of exemplary ukemi techniques and to obtain kinematic information of the head, in a sequence of ukemi from the onset of the throw to the completion of ukemi. The results indicated that the vertical velocity with which the uke's head decelerated was reduced by increasing the body surface exposed to the collision with the tatami and by increasing the elapsed time. In particular, overall upper limb contact with the tatami is greatly associated with deceleration. In o-soto-gari, the impulsive force on the faller's head as the head reached the lowest point was 204.82 ± 19.95 kg m · s(-2) while in o-uchi-gari it was 118.46 ± 63.62 kg m · s(-2), z = -1.75, P = 0.08, and it did present a large-sized effect with r = 0.78. These findings indicate that the exemplary o-soto-gari as compared to o-uchi-gari is the technique that causes more significant damage to the uke's head.

Combined treatment with bortezomib plus bafilomycin A1 enhances the cytocidal effect and induces endoplasmic reticulum stress in U266 myeloma cells: crosstalk among proteasome, autophagy-lysosome and ER stress.

Bortezomib (BZ), a first line 26S proteasome inhibitor, induces a potent cytocidal effect with caspase-3 activation in multiple myeloma (MM) cell lines. Since IκBα is a substrate of the proteasome, the initial rationale for using BZ in MM has been to inhibit NF-κB. However, BZ rather activated NF-κB activity in U266 cells. BZ induces autophagy as well as endoplasmic reticulum (ER) stress in various cell lines tested. Inhibition of initial autophagosome formation by treatment with either 3-methyladenine or siRNA for LC3B in U266 cells and knockdown of the atg5 gene in a murine embryonic fibroblastic cell line all resulted in attenuation of BZ-induced cell death. In contrast, combined treatment with BZ and bafilomycin A1 (BAF), which is a specific inhibitor of vacuolar-ATPase and is used as an autophagy inhibitor at the late stage, resulted in synergistic cytotoxicity, compared with that by either BZ or BAF alone. BAF treatment also induced ER stress, but the kinetics of inductions of ER stress-related genes [e.g. CHOP (GADD153) and GRP78] completely differed between BZ- and BAF-treatments: BZ induced these ER stress markers within 8 h, whereas treatment with BAF required more than 48 h in U266 cells. In order to synchronize ER stress, we pre-treated U266 cells with BAF for 48 h, followed with BZ for 48 h. The sequential treatment with BAF and BZ induced a further enhanced cytotoxicity, compared with the simultaneous combination of BAF and BZ. These data suggest crosstalk among the ubiquitin-proteasome system, the autophagy-lysosome system, and ER stress. Controlling these interactions and kinetics appears to have important implications for optimizing clinical cancer treatment including MM-therapy.

Isoflavones inhibit nicotine C-oxidation catalyzed by human CYP2A6.

The inhibitory effects of isoflavones (daidzein, genistein, and glycitein) on human cytochrome P450 (CYP) 2A6 activities were investigated. Daidzein, genistein, and glycitein uncompetitively inhibited nicotine C-oxidation catalyzed by recombinant CYP2A6 expressed in baculovirus-infected insect cells with Ki values of 1.3 +/- 0.3 microM, 0.7 +/- 0.2 microM, and 5.2 +/- 0.8 microM, respectively, but not coumarin 7-hydroxylation. Effects of the intake of soy isoflavones on in vivo nicotine metabolism were investigated with 7 healthy Japanese homozygotes of CYP2A6*1. The cotinine/nicotine ratio of the plasma concentrations 2 hours after chewing 1 piece of nicotine gum under the basal condition (after abstaining from soy foods for 1 week) was 8.8 +/- 2.6 (4.4-11.4). The ratio was significantly (P < .05) reduced to 6.7 +/- 1.6 (4.0-8.2) after consumption of a soy isoflavone supplement (60 mg of total isoflavones/d) for 5 days. The authors found that isoflavone contained in soy products significantly decreased nicotine metabolism.

Di-(2-ethylhexyl) phthalate induces severe aspermatogenesis in mice, however, subsequent antioxidant vitamins supplementation accelerates regeneration of the seminiferous epithelium.

Di-(2-ethylhexyl) phthalate (DEHP), now regarded as an endocrine disruptor, can experimentally induce spermatogenic disturbance in laboratory animals. Our previous study demonstrated that antioxidant vitamins (vitamins C and E) supplementation during DEHP-treatment significantly protected the rat seminiferous epithelium from DEHP-gonadotoxicity. In the present study, we gave these antioxidant vitamins to mice already having fully developed aspermatogenesis because of DEHP to determine whether or not the vitamins can cure the injured seminiferous epithelium. CD-1 male mice were fed on a DEHP-containing diet for 15 days and then fed on the DEHP-free normal diet with or without supplementation of vitamins C and E in drinking water for another 50 days. The results showed that severe aspermatogenesis was induced by the DEHP-treatment but that the damaged seminiferous epithelium spontaneously recovered whether the vitamins were provided or not. This indicates that the DEHP-induced aspermatogenesis was reversible. However, the supplementation of antioxidant vitamins significantly accelerated regeneration of the injured seminiferous epithelium, suggesting that the vitamins have a therapeutic effect on DEHP-induced aspermatogenesis.

Toluene diisocyanate exposure induces laryngo-tracheal eosinophilia, which can be ameliorated by supplementation with antioxidant vitamins in guinea pigs.

OBJECTIVE: Toluene diisocyanate (TDI)-induced asthma is a common cause of occupational lung disease. In addition, a sore throat is one of the complaints of TDI-exposed workers. The aim of the present study was to determine whether TDI exposure induces laryngeal and/or tracheal lesions in experimental animals. MATERIAL AND METHODS: Guinea pigs underwent naris application of TDI three times, and their respiratory tracts were then examined using light and electron microscopy. Some animals simultaneously received vitamins C and E. which function as antioxidant agents. RESULTS: When TDI-treated animals showed the clinical sign of labored breathing, many eosinophils had appeared in the lamina propria and mucosa of both the larynx and trachea, which finally infiltrated the tract lumen through the ruptured epithelium. Laryngo-tracheal inflammation was more severe than that observed in the lungs. However, supplementation with antioxidant vitamins in TDI-treated animals ameliorated the respiratory eosinophilia. CONCLUSION: Naris application of TDI induced laryngotracheitis. which was significantly suppressed by the antioxidant vitamins, This implies a preventive effect of the vitamins on this occupational disease.