Takehiko Nakamura

Pars plana vitrectomy assisted by triamcinolone acetonide for refractory uveitis: a case series study

Aim: To examine the outcome of a triamcinolone acetonide (TA) assisted pars plana vitrectomy (PPV) for refractory uveitis. Methods: Six patients suffering from proliferative vitreoretinopathy (PVR) with refractory uveitis underwent a TA assisted PPV. The patients consisted of one with Vogt-Koyanagi-Harada disease, one with acute retinal necrosis, one with Behçet’s disease, and three with sarcoidosis. TA was inoculated into the vitreous cavity to visualise the vitreous. In four of six patients, 4 mg of TA were intentionally left in the vitreous cavity to reduce the degree of postoperative inflammation. Results: The vitreous body was clearly seen using TA during surgery, which greatly helped us to perform a posterior hyaloid resection safely and thoroughly. As we previously observed in other disease, TA allowed us to visualise the transparent vitreous and thus was helpful in removing the vitreous cortex from the retina completely in uveitis. One patient (Behçet’s disease, in whom TA was intentionally left) showed an elevated intraocular pressure (IOP) transiently after surgery which was controllable by topical eye drops. The remaining TA diminished day by day and had almost completely disappeared within a month from operation. Conclusion: TA improved the visibility of the hyaloid and the safety of the surgical procedures and no serious complications were observed after TA assisted PPV in uveitis. Although the long term effects are still unknown, this method appears to be potentially useful as an improved treatment for PVR associated with refractory uveitis.

The protective role of T-cell receptor Vγ1+ T cells in primary infection with Listeria monocytogenes

We have previously reported that the T‐cell receptor (TCR) γδ+ T cells increase in mice infected with an intracellular bacteria Listeria monocytogenes, and the cells predominantly express Vδ6 and Vγ1 genes. In this study, we used a monoclonal antibody (mAb) specific to TCR Vγ1 to estimate the frequency of Vγ1+ T cells and we discuss their significance in protection against L. monocytogenes. The spleen, liver and peritoneal exudate cells from mice intraperitoneally infected with L. monocytogenes were analysed by flow cytometry. In all the organs investigated, Vγ1+ cells increased predominantly among TCR γδ+ T cells at an early phase (day 5–7) of the infection. To elucidate the significance of the Vγ1+ T cells in the protection against L. monocytogenes, mice were depleted of TCR Vγ1+γδ T cells or all TCR γδ+ T cells by intraperitoneal inoculation of anti‐Vγ1 mAb or anti‐pan TCR γδ mAb, respectively, before infection with L. monocytogenes. The bacterial growth in the spleen and the liver examined on day 5 after the infection increased significantly by the depletion of TCR Vγ1+ T cells. The numbers of L. monocytogenes in TCR Vγ1+ T‐cell‐depleted mice were nearly the same as in mice depleted of all TCR γδ+ T cells. These results demonstrated that Vγ1+ T cells are the predominant population of γδ T cells in protection against L. monocytogenes at the early phase of the primary infection.

The role of γδ T cells in induction of bacterial antigen‐specific protective CD8+ cytotoxic T cells in immune response against the intracellular bacteria Listeria monocytogenes

The role of T‐cell receptor (TCR) γδ T cells in the induction of protective TCR αβ T cells against infection by the intracellular bacteria Listeria monocytogenes was analysed. We found that depletion of γδ T cells by anti‐TCR δ monoclonal antibody treatment before intravenous immunization of mice with a sublethal dose of viable L. monocytogenes resulted in reduction of protection against secondary challenge infection in the immunized mice. The γδ T‐cell depletion also reduced induction of protective αβ T cells capable of transferring the protection against challenge infection of L. monocytogenes into naive mice. Furthermore, the protective T cells that were affected by the γδ T‐cell depletion were suggested to be CD8+ cytotoxic T cells rather than CD4+ T cells by the following observations. First, induction of cytotoxic T lymphocytes specific to a L. monocytogenes‐derived H‐2Kd‐restricted peptide (listeriolysin O 91–99) was significantly suppressed by γδ T‐cell depletion before immunization. Second, γδ T‐cell depletion did not affect cytokine production and proliferation of T cells from immunized mice in response to in vitro stimulation with heat‐killed Listeria which preferentially stimulates CD4+ T cells. Third, CD8+αβ T cells from control immunized mice transferred protection against infection of L. monocytogenes into naive mice but only a limited degree of protection was transferred by CD8+ T cells from the γδ T‐cell‐depleted immunized mice; and fourth, CD4+αβ T cells from the γδ T‐cell‐depleted mice transferred a similar level of protection as those from the control immunized mice. All these results suggest that γδ T cells participate in establishment of protective immunity against intracellular bacteria by supporting priming of bacterial antigen‐specific CD8+ cytotoxic T cells.

Persistent infection with Listeria monocytogenes in the kidney induces anti-inflammatory invariant fetal-type γδ T cells

After intraperitoneal inoculation with Listeria monocytogenes, γδ T cells appear in the peritoneal cavity preceding the appearance of αβ T cells. Such γδ T cells predominantly express T‐cell receptor (TCR)Vγ1/Vδ6, develop through an extrathymic pathway, and contribute to host defence against the bacteria. We have observed a gradual increase in γδ T cells in kidneys of mice after intrarenal inoculation with L. monocytogenes, which resulted in an unusually long‐lasting local infection. In this study, we examined the characteristics and the roles of the γδ T cells induced in this model. It was found that these γδ T cells predominantly expressed TCRVγ6/Vδ1 with canonical junctional sequences identical to those expressed on fetal thymocytes. Although depletion of such γδ T cells in vivo did not affect the number of bacteria, it resulted in histologically exacerbated inflammation in the kidneys. These results indicate that a persistent infection with L. monocytogenes in kidneys induces a different kind of γδ T cell from that induced after intraperitoneal infection. The former expresses invariant fetal‐type Vγ6/Vδ1+TCR and plays a regulatory role in resolution of inflammation.

The comparative benefits of glaucoma filtering surgery with an electric-pulse targeted drug delivery system demonstrated in an animal model

PURPOSE To evaluate the safety and effectiveness of glaucoma filtering surgery performed with the adjunctive use of bleomycin administered in conjunction with electric pulses (EP). DESIGN Experimental study in rabbits. CONTROLS AND METHODS: Trabeculectomies were performed on pigmented rabbits (2 to 2.5 kg) using the following adjunctive treatments: 5 microM of topical bleomycin and EP (5V, 50 msec, 8 pulses) (group A: B+E+, n=15); bleomycin but no EP (group B: B+E-, n=15); 5 ,uM mitomycin C (MMC) and EP (group C: M+E+, n= 10); MMC but no EP (group D: M+E-, n=10); EP alone (group E: E+, n=10); and no adjunctive treatment (group F: E-, negative control, n=10). MAIN OUTCOME MEASURES Intraocular pressure (IOP) was measured regularly for 60 days after the operation. Bleb formation and the condition of the conjunctiva, cornea, and retina were also regularly evaluated. Histologic studies were performed by light microscopy, and retinal functions were evaluated by electroretinography. RESULTS Postoperative IOP was significantly lower than the preoperative level in all the animals until day 7. However, in groups E and F (the negative control) it returned to the preoperative level after day 7, and in groups B, C, and D after 15 days. The IOP of group A remained lower even on day 40. The average amount IOP was lowered or increased on day 20 was -6.4 mmHg (P < 0.05) in group A; -0.2 mmHg in group B; +1.2 mmHg in group C; and -3.25 mmHg in group D. The survival rate of the filtering blebs on day 20 was significantly higher in group A than in the other groups. Clinical and histologic studies uncovered no pathologic findings in any intra- or paraocular tissues. Electroretinographic evaluation of retinal function in group A showed no apparent change over the 60 days of the study. CONCLUSION Glaucoma filtering surgery in rabbits with the adjunctive use of bleomycin in conjunction with EP significantly lowered IOP for up to 40 days without clinically, morphologically, or functionally harming intraocular tissues.

THE CHARACTERIZATION OF TESTICULAR CELL (TC)-SPECIFIC T-CELL CLONES INDUCED BY INTRATESTICULAR LISTERIA MONOCYTOGENES INFECTION : TC-SPECIFIC T CELLS WITH ATYPICAL CYTOKINE PROFILE TRANSFER ORCHITIS

A unilateral infection of Listeria monocytogenes into the testis of mice induces not only Listeria‐specific T cells but also autoreactive T cells that can transfer experimental autoimmune orchitis (EAO) into naive mice. To investigate the characteristics of the autoreactive T cells, we established six testicular cell (TC)‐specific T‐cell clones from the spleen of the intratesticularly infected mice. All the clones expressed CD4 and T‐cell receptor (TCR) αβ, and four of the six clones expressed Vβ8. They showed proliferative response to TC in the presence of syngeneic spleen antigen‐presenting cells, but did not cross‐react to Listeria antigen (Ag). They produced interferon‐γ (IFN‐γ) when stimulated with TC, but interleukin‐2 (IL‐2), IL‐4 and IL‐10 were undetectable. IL‐2 production was not detected even when they were restimulated with TC after a 10‐day resting culture without Ag and IL‐2, although they proliferated in the restimulation culture. Even in the presence of anti‐IL‐2 mAb, the TC‐specific T‐cell clones showed proliferative response against TC. The observations indicate that the TC‐specific IFN‐γ‐producing T cells proliferate in the absence of autocrine. Both intravenous and intratesticular injection of these clones transferred EAO in syngeneic naive mice. These results suggest that L. monocytogenes infection in the testis induces autoreactive orchitogenic CD4+ T cells without cross‐reactivity to bacterial Ag. Furthermore, these data demonstrate that CD4+ T cells with an atypical cytokine profile can efficiently cause EAO.

Macrophages activated by Listeria monocytogenes induce organ-specific autoimmunity.

We have previously reported an experimental autoimmune model induced by the local infection of Listeria monocytogenes. The unilateral inoculation of virulent Listeria into a testis of a normal mouse induced a delayed‐type hypersensitivity response against testicular antigen and caused autoimmune orchitis in the contralateral testis. The orchitis was transferred to naive mice by T cells from the intratesticularly infected mice. In this paper, we demonstrated that avirulent Listeria, which lacks the expression of listeriolysin O, failed to induce any anti‐testicular responses or contralateral orchitis even when it was inoculated at a high dose into the testis. Furthermore, the intraperitoneal inoculation of virulent Listeria with testicular antigen induced the anti‐testicular responses and orchitis although intraperitoneal inoculation of testicular antigen with avirulent Listeria failed to induce them. The difference between virulent and avirulent Listeria in the induction of anti‐testicular responses was supposed to be dependent on the difference in macrophage activation by the two bacterial strains because, first, the anti‐testicular responses were elicited in normal mice when macrophages from virulent Listeria‐infected mice were intraperitoneally transferred with testicular antigen although no viable bacteria were detected from the macrophages, and secondly, in contrast, the intraperitoneal co‐inoculation of macrophages from avirulent Listeria‐infected mice and testicular antigen failed to elicit any anti‐testicular responses. Finally, we found that the virulent Listeria‐induced macrophages expressed a higher level of CD80 (B7‐1) and CD86 (B7‐2) molecules than did the avirulent Listeria‐induced macrophages and naive peritoneal macrophages. These results thus suggest that virulent Listeria activates macrophages to induce autoreactive T cells while avirulent Listeria does not. The up‐regulation of B7 molecules by virulent Listeria infection is a candidate of the mechanism for the activation of autoreactive T cells.

Absolute Calibration by Helium-Ion-Implanted Samples for Helium Accumulation Method Applied to Neutron Dosimetry

A He accumulation method applied to neutron dosimetry is a useful means to measure a fast-neutron fluence in a high-flux neutron field. The number of He atoms produced by (n, xα) reactions should be absolutely measured in this method. One of the methods of the calibration of a He atom measurement system is a standard He gas method which is based on the measurement of the pressure, volume and temperature of the He gas. The uncertainty in measuring the number of He atoms is largely influenced by the standard gas purity. In this paper, a He-ion-implanted sample method has been developed as a new method for the calibration. The number of implanted He ions can be determined by measuring the current of ions analyzed by a magnetic field where uncertainty of the measurement can be made reasonably small as it depends only on the ion current measurement. The two methods have been compared by using Al samples implanted with He ions. Both the methods can be applied as a mutually complementary calibration each. The He...

He 集積法による中性子照射量測定のための He 絶対量測定装置

In the helium accumulation method for neutron dosimetry, it is applied that the number of helium atoms produced through (n,xα) reaction is proportional to neutron fluence in fast neutron irradiation fields. Besides, activities measuring in the activation method saturate to the value depending on their half-lives. A helium atom measurement, system for detecting of helium atoms accumulated in neutron-irradiated samples has been constructed and tested. The samples are vaporized in a furnace to release helium contained in them. The number of helium atoms is measured with a quadrupole mass spectrometer calibrated by a standard helium gas supply which is consist of a known-volume vessel, a pressure gauge and a thermometer. It has been proposed in this work that helium-ion implanted aluminum disks are applied to calibrate the helium atom measurement system. This method has succeeded to obtain results agreed with the standard helium gas supply system.