Naoyuki Hatayama

Hydrogen-rich University of Wisconsin solution attenuates renal cold ischemia-reperfusion injury.

Background Renal ischemia-reperfusion (I/R) injury is unavoidable in kidney transplantation and frequently influences both short- and long-term allograft survival rates. One of the major events in I/R injury is the generation of cytotoxic oxygen radicals. Recently, hydrogen gas has been reported to display antioxidant properties and protective effects against organ dysfunction induced by various I/R injuries. We investigated whether hydrogen-rich University of Wisconsin (HRUW) solution attenuates renal cold I/R injury. Methods We prepared HRUW solution by a novel method involving immersion of centrifuge tubes containing UW solution into hydrogen-saturated water. Hydrogen readily permeates through the centrifuge tubes, and thus, the hydrogen concentration of the UW solution gradually increases in a time-dependent manner. Syngeneic rat kidney transplantation was performed, and the animals were divided into three groups: recipients with nonpreserved grafts (control group), recipients with grafts preserved in UW solution for 24 to 48 hr (UW group), and recipients with grafts preserved in HRUW solution for 24 to 48 hr (HRUW group). Results In the early phases, HRUW solution decreased oxidative stress, tubular apoptosis, and interstitial macrophage infiltration in the kidney grafts. Consequently, HRUW solution improved renal function and prolonged recipient survival rate compared with simple cold storage using UW solution. Histopathologically, HRUW treatment alleviated tubular injury and suppressed development of interstitial fibrosis. Conclusions HRUW solution improved graft function and prolonged graft survival compared with simple cold storage using UW solution by protecting tubular epithelial cells from inflammation and apoptosis. Our new method of organ preservation is a groundbreaking, safe, and simple strategy that may be applied in the clinical setting.

History and future of human cadaver preservation for surgical training: from formalin to saturated salt solution method

Traditionally, surgical training meant on-the-job training with live patients in an operating room. However, due to advancing surgical techniques, such as minimally invasive surgery, and increasing safety demands during procedures, human cadavers have been used for surgical training. When considering the use of human cadavers for surgical training, one of the most important factors is their preservation. In this review, we summarize four preservation methods: fresh-frozen cadaver, formalin, Thiel’s, and saturated salt solution methods. Fresh-frozen cadaver is currently the model that is closest to reality, but it also presents myriad problems, including the requirement of freezers for storage, limited work time because of rapid putrefaction, and risk of infection. Formalin is still used ubiquitously due to its low cost and wide availability, but it is not ideal because formaldehyde has an adverse health effect and formalin-embalmed cadavers do not exhibit many of the qualities of living organs. Thiel’s method results in soft and flexible cadavers with almost natural colors, and Thiel-embalmed cadavers have been appraised widely in various medical disciplines. However, Thiel’s method is relatively expensive and technically complicated. In addition, Thiel-embalmed cadavers have a limited dissection time. The saturated salt solution method is simple, carries a low risk of infection, and is relatively low cost. Although more research is needed, this method seems to be sufficiently useful for surgical training and has noteworthy features that expand the capability of clinical training. The saturated salt solution method will contribute to a wider use of cadavers for surgical training.

Contribution of IL-12/IL-35 common subunit p35 to maintaining the testicular immune privilege.

The testis is an organ with immune privilege. The comprehensive blood–testis barrier formed by Sertoli cells protects autoimmunogenic spermatozoa and spermatids from attack by the body’s immune system. The interleukin (IL)-6/IL-12 family cytokines IL-12 (p35/p40), IL-23 (p19/p40), IL-27 (p28/Epstein-Barr virus−induced gene 3 [EBI3]), and IL-35 (p35/EBI3) play critical roles in the regulation of various immune responses, but their roles in testicular immune privilege are not well understood. In the present study, we investigated whether these cytokines are expressed in the testes and whether they function in the testicular immune privilege by using mice deficient in their subunits. Expression of EBI3 was markedly increased at both mRNA and protein levels in the testes of 10- or 12-week-old wild-type mice as compared with levels in 2-week-old mice, whereas the mRNA expression of p40 was markedly decreased and that of p35 was conserved between these two groups. Lack of EBI3, p35, and IL-12 receptor β2 caused enhanced infiltration of lymphocytes into the testicular interstitium, with increased interferon-γ expression in the testes and autoantibody production against mainly acrosomal regions of spermatids. Spermatogenic disturbance was more frequently observed in the seminiferous tubules, especially when surrounded by infiltrating lymphocytes, of these deficient mice than in those of wild-type mice. In particular, p35-deficient mice showed the most severe spermatogenic disturbance. Immunohistochemical analyses revealed that endothelial cells and peritubular cells in the interstitium were highly positive for p35 at both ages, and CD163+ resident macrophages positive for p35 and EBI3, possibly producing IL-35, were also detected in the interstitium of 12-week-old mice but not those of 2-week-old mice. These results suggest that p35 helps in maintaining the testicular immune privilege, in part in an IL-35-dependent manner.

Saturated Salt Solution Method: A Useful Cadaver Embalming for Surgical Skills Training

AbstractThis article evaluates the suitability of cadavers embalmed by the saturated salt solution (SSS) method for surgical skills training (SST).SST courses using cadavers have been performed to advance a surgeons techniques without any risk to patients. One important factor for improving SST is the suitability of specimens, which depends on the embalming method. In addition, the infectious risk and cost involved in using cadavers are problems that need to be solved.Six cadavers were embalmed by 3 methods: formalin solution, Thiel solution (TS), and SSS methods. Bacterial and fungal culture tests and measurement of ranges of motion were conducted for each cadaver. Fourteen surgeons evaluated the 3 embalming methods and 9 SST instructors (7 trauma surgeons and 2 orthopedists) operated the cadavers by 21 procedures. In addition, ultrasonography, central venous catheterization, and incision with cauterization followed by autosuture stapling were performed in some cadavers.The SSS method had a sufficient antibiotic effect and produced cadavers with flexible joints and a high tissue quality suitable for SST. The surgeons evaluated the cadavers embalmed by the SSS method to be highly equal to those embalmed by the TS method. Ultrasound images were clear in the cadavers embalmed by both the methods. Central venous catheterization could be performed in a cadaver embalmed by the SSS method and then be affirmed by x-ray. Lungs and intestines could be incised with cauterization and autosuture stapling in the cadavers embalmed by TS and SSS methods.Cadavers embalmed by the SSS method are sufficiently useful for SST. This method is simple, carries a low infectious risk, and is relatively of low cost, enabling a wider use of cadavers for SST.

Effect of a Magnetic Field on Drosophila under Supercooled Conditions

Under subzero degree conditions, free water contained in biological cells tends to freeze and then most living things die due to low temperatures. We examined the effect of a variable magnetic field on Drosophila under supercooled conditions (a state in which freezing is not caused even below the freezing point). Under such supercooled conditions with the magnetic field at 0°C for 72 hours, −4°C for 24 hours and −8°C for 1 hour, the Drosophila all survived, while all conversely died under the supercooled conditions without the magnetic field. This result indicates a possibility that the magnetic field can reduce cell damage caused due to low temperatures in living things.

Effects on the local immunity in the testis by exposure to di-(2-ethylhexyl) phthalate (DEHP) in mice.

Exposure to di-(2-ethylhexyl) phthalate (DEHP) has been reported to induce spermatogenic disturbance through oxidant stress and affect the immune system as an adjuvant. However, the effect of DEHP on the testicular immune microenvironment has not yet been investigated. In the present study, we examined the testicular immune microenvironment after exposure to doses of DEHP, previously identified as no-observed-adverse-effect levels. Adult male mice were administered food containing 0%, 0.01% or 0.1% DEHP and then testes were analyzed. The results showed that a slight but significant spermatogenic disturbance appeared in the 0.1% DEHP group but not in the 0.01% DEHP group at 8 weeks. It was also demonstrated that lymphocytes and F4/80- and MHC class II- positive cells were significantly increased with the elevation of IL-10 and IFN-γ mRNA expressions in the testes of not only the 0.1% DEHP group but also the 0.01% DEHP group at 8 weeks. Histochemical analyses involving horseradish peroxidase (HRP) as a tracer showed that a little blood-borne HRP had infiltrated into the lumen of a few seminiferous tubules beyond the blood-testis-barrier in both the 0.1% and 0.01% DEHP groups at 8 weeks. This indicates that a dose of DEHP that has little effects on spermatogenesis can change the testicular immune microenvironment with functional damage of the blood-testis barrier.

Low-dose exposure to di-(2-ethylhexyl) phthalate (DEHP) increases susceptibility to testicular autoimmunity in mice

Exposure to di-(2-ethylhexyl) phthalate (DEHP) induces spermatogenic disturbance (SD) through oxidative stress, and affects the immune system by acting as an adjuvant. Recently, we reported that in mice, a low dose of DEHP, which did not affect spermatogenesis, was able to alter the testicular immune microenvironment. Experimental autoimmune orchitis (EAO) can be induced by repeated immunization with testicular antigens, and its pathology is characterized by production of autoantibodies and SD. In the present study, we investigated the effect of a low-dose DEHP on the susceptibility of mice to EAO. The exposure to DEHP-containing feed (0.01%) caused a modest functional damage to the blood-testis barrier (BTB) with an increase in testicular number of interferon gamma (IFN-γ)-positive cells and resulted in the production of autoantibodies targeting haploid cells, but did not affect spermatogenesis. While only single immunization with testicular antigens caused very mild EAO, the concurrent DEHP exposure induced severe EAO with significant increases in number of interferon gamma-positive cells and macrophages, as well as lymphocytic infiltration and serum autoantibody titer accompanied by severe SD. To summarize, the exposure of mice to the low-dose DEHP does not induce significant SD, but it may cause an increase in IFN-γ positive cells and modest functional damage to the BTB in the testis. These changes lead to an autoimmune response against haploid cell autoantigens, resulting in increased susceptibility to EAO.

The Effects of Adjuvants on Autoimmune Responses Against Testicular Antigens in Mice

Abstract Experimental autoimmune orchitis (EAO) is a model of immunologic male infertility and pathologically characterized by lymphocytic inflammation, which causes breakdown of the testicular immune privilege with spermatogenic disturbance. Generally, murine EAO is induced by immunization with testicular homogenate (TH) from the testes of donor mice + complete Freunds adjuvant (CFA) + Bordetella pertussigens (BP), and it has been considered that treatment with these two adjuvants is required to enhance the immune response against testicular antigens. However, there remains a possibility that CFA and BP may affect autoimmune responses against the testicular antigens without TH. In the present study, we examined this possibility using real-time RT-PCR, Western blotting and immunohistochemical staining. The results demonstrated that immunization with TH in combination with CFA and BP evoked more severe EAO than that with only TH. Real-time RT-PCR analyses revealed that Fas mRNA expression in TH+CFA+BP-induced EAO was significantly higher than that in TH-induced EAO. Interestingly, IL-6 mRNA expression dramatically increased in TH+CFA+BP-induced EAO; however, no apparent change in IL-6 mRNA expression occurred in TH-induced EAO. It was also noted that treatment with CFA and BP alone augmented autoimmune reactions against some testicular autoantigens. These results indicates that these adjuvants are helpful in evoking severe EAO, and treatment with the adjuvants alone can evoke autoimmune reactions against some testicular autoantigens despite the use of no TH.

Serum Autoantibodies in Mice Immunized with Syngeneic Testicular Germ Cells Alone

The aim of this study was to identify the characteristics of serum autoantibodies in experimental autoimmune orchitis (EAO) induced by immunization with syngeneic testicular germ cells (TGC) alone in mice.

Preservation by desiccation of isolated rat hearts for 48 hours using carbon monoxide (PCO = 4,000 hPa) and oxygen (PO(2) = 3,000 hPa).

It is currently said that CO has anti-inflammatory and antiapoptosis effects and it has attracted attention as a medical gas. We used CO for rat hearts and conducted a preservation experiment. We isolated rat hearts, placed them into a specially made chamber, filled the chamber with a gas mixture of PCO (4,000 hPa) and PO2 (3,000 hPa), and preserved the hearts in a refrigerator at 4°C for 48 h. We then performed a heterotrophic transplantation on the neck of each recipient rat and resuscitated the preserved hearts. We herein report our findings.