Two-signal blockade with anti-CD45RB and anti-CD154 monoclonal antibodies inhibits graft rejection via CD4-dependent mechanisms in allogeneic skin transplantation

Blockade of signal 1 or 2 for T-cell activation by the use of anti-CD45RB and anti-CD154 monoclonal antibodies (mAb) (two-signal blockade) has been proven effective in preventing or delaying graft rejection. However, the mechanisms of its immunomodulatory effects are clearly unknown and the present studies were performed to determine how the two-signal blockade modulate allogeneic immune responses, especially T-cell mediated cellular immunity, in a murine skin allograft model. We now report on the profound inhibition of alloreactive T cells by two-signal blockade via CD4-dependent mechanisms. C57BL/6 mice of BALB/c skin allograft were treated with anti-CD45RB, anti-CD154, CTLA4-Ig, or their combinations. For depletion of CD4 or CD8 T cells, the recipients received CD4-depleting or CD8-depleting mAb. We confirmed that survival of skin allograft was markedly prolongated in the two-signal blockade-treated group. In depletion study, anti-CD45RB, anti-CD154 and CD4-depleting mAb-treated group showed acute rejection of skin allograft in contrast to CD8-depleting group treated with the two-signal blockade. In the group treated with the two-signal blockade, the proportions of CD4+CD45RB(low) and CD8+CTLA-4 regulatory T cells were increased while effector CD8+ T cells, including IFN-gamma-secreting and CD8+CD62L(low) T cells, were decreased when compared with non-treated group. In contrast, the CD4-depleted group treated with the two-signal blockade resulted in recovery from immunoregulatory effects of two-signal blockade. In addition, results of IL-4 and IL-10 production were also showed CD4-dependence. Therefore, the two- signal blockade is accompanied by CD4-dependent mechanisms in allogeneic skin transplantation.     

4-1BB (CD137) signals depend upon CD28 signals in alloimmune responses

Our previous study has demonstrated that there is a significant delay of Balb/c cardiac allograft rejection in the C57BL/6 4-1BB-deficient knockout recipient. In this study, we examined the effect of combined blockade of the 4-1BB and CD28 costimulatory pathways on cardiac allograft rejection in the C57BL/6-->Balb/c model. A long-term cardiac allograft survival was induced in CD28/4-1BB- deficient mice (>100 days survival in 3 of 4 mice), which was comparable with CD28-deficient mice (>100 days survival in 2 of 5 mice; P<0.2026). There was no long-term cardiac allograft survival in either wild-type (WT) or 4-1BB-deficient mice, even though 4-1BB-deficient recipients showed a significant delay of cardiac allograft rejection than WT mice. An in vitro mixed leukocyte reaction (MLR) assay showed that 4-1BB-deficient and WT mouse T cells had a similar responsiveness to allostimulation, whereas CD28- and CD28/4-1BB-deficient mouse T cells had a defective responsiveness to allostimulation. Furthermore, 4-1BB-deficient mice showed a similar CTL but an elevated Ab response against alloantigens as compared to WT mice, and the alloimmune responses of 4-1BB-deficient mice were abrogated in the CD28-deficient background. Overall, these results indicate that the CD28 costimulatory pathway plays a major role in the alloimmune response and that 4-1BB signals are dependent upon CD28 signals.     

Enhancement of cutaneous immune response to bacterial infection after low-level light therapy with 1072 nm infrared light: a preliminary study

We investigated the photobiomodulation effects of 1072 nm infrared light on the natural immune response involved in anti-bacterial and wound healing processes. Thirty mice infected with MRSA on the skin were divided into two groups. The experimental group was treated with 1072 nm infrared light (irradiance: 20 mW/cm(2), fluence: 12 J/cm(2) for 10 min) at 2, 4, 8, 12, 24 h, 3 and 5 days after inoculation and the control group with sham light. Serial changes of the mRNA levels of TLR2, IL-1β, TNF-α, IL-6, iNOS, MCP-1, TGF-β, bFGF and VEGF were studied by real time RT-PCR and those of the expression level of VEGF, bFGF, TGF-β and NF-κB by immunohistochemistry. The mRNA levels of the cytokines involved in the early phase of anti-bacterial immune response (IL-1β, TNF-α, IL-6, MCP-1) increased significantly in the 1072 nm group, peaking between 12 and 24 h post-inoculation. These levels normalized after 3-5 days. Immunohistochemistry revealed a notably stronger expression of VEGF in the 1072 nm group from 8-h post-inoculation to 5-day post-inoculation. We concluded that 1072 nm infrared light had a photobiomodulation effect which resulted in an enhanced biological immune response to the bacterial infection by MRSA and also increased the expression of VEGF to a significant level.     

The effects of grape seeds polyphenols on SKH-1 mice skin irradiated with multiple doses of UV-B

The study investigated the protective activity of red grape seeds (Vitis vinifera L, Burgund Mare variety) (BM) extracts in vivo on multiple doses of ultraviolet radiation (UV)-B-induced deleterious effects in SKH-1 mice skin. Eighty 8-weeks-old female SKH-1 mice were divided into 8 groups: control, vehicle, UV-B irradiated, vehicle+UV-B irradiated, BM 2.5mg polyphenols (PF)/cm(2)+UV-B irradiated, BM 4 mg PF/cm(2)+UV-B irradiated, UV-B+BM 2.5mg PF/cm(2), UV-B+BM 4 mg PF/cm(2). The extract was applied topically before or after each UV-B exposure (240 mJ/cm(2)), for 10 days consecutively. The antioxidant activity of BM extract is higher than gallic acid (k(BM)=0.017, k(gallic acid)=0.013). Multiple doses of UV-B generated the formation of cyclobutane pyrimidine dimers (CPDs) and sunburn cells, increased glutathione peroxidase (GPx) and catalase (CAT) activities respectively glutathione (GSH) and IL-1β levels in skin. In group treated with 2.5mg PF/cm(2) before UV-B irradiation BM extract inhibited UV-B-induced sunburn cells, restored the superoxide dismutase (MnSOD) activity, increased insignificantly CAT and GPx activities and reduced IL-1β level. The BM 4.0 mg PF/cm(2) treatment decreased GSH level and reduced the percentage of CPDs positive cells in skin. Both doses of BM extract administered after UV-B irradiation increased the MnSOD and GPx activities and reduced the formation of sunburn cells in skin. Our results suggest that BM extract might be a potential chemo-preventive candidate in reducing the oxidative stress and apoptosis induced by multiple doses of UV-B in skin.     

Bilirubin protects grafts against nonspecific inflammation-induced injury in syngeneic intraportal islet transplantation

Nonspecific inflammatory response is the major cause for failure of islet grafts at the early phase of intraportal islet transplantation (IPIT). Bilirubin, a natural product of heme catabolism, has displayed anti-oxidative and anti-inflammatory activities. The present study has demonstrated that bilirubin protected islet grafts by inhibiting nonspecific inflammatory response in a syngeneic rat model of IPIT. The inflammation-induced cell injury was mimicked by exposing cultured rat insulinoma INS-1 cells to cytokines (IL-1β, TNF-α and IFN-γ) in in vitro assays. At appropriate lower concentrations, bilirubin significantly attenuated the reduced cell viability and enhanced cell apoptosis induced by cytokines, and protected the insulin secretory function of INS-1 cells. Diabetic inbred male Lewis rats induced by streptozotocin underwent IPIT at different islet equivalents (IEQs) (optimal dose of 1000, and suboptimal doses of 750 or 500), and bilirubin was administered to the recipients every 12 h, starting from one day before transplantation until 5 days after transplantation. Administration of bilirubin improved glucose control and enhanced glucose tolerance in diabetic recipients, and reduced the serum levels of inflammatory mediators including IL-1β, TNF-α, soluble intercellular adhesion molecule 1, monocyte chemoattractant protein-1 and NO, and inhibited the infiltration of Kupffer cells into the islet grafts, and restored insulin-producing ability of transplanted islets.     

DNA ASSOCIATED WITH THE CELL MEMBRANE IS INVOLVED IN THE INHIBITION OF THE SKIN REJECTION RESPONSE INDUCED BY INFUSIONS OF PHOTODAMAGED ALLOREACTIVE CELLS THAT MEDIATE REJECTION OF SKIN ALLOGRAFT

Abstract Cell membrane DNA (cmDNA) is a form of DNA located on the surface of human and murine T-cells. It has recently been characterized as a target for photomodification by 8-methoxypsoralen (8-MOP) and long-wave ultraviolet light (UV-A). Whereas 8-MOP itself is biologically inert, photoactivated 8-MOP is covalently bound to pyrimidine bases in DNA. We have investigated the possible involvement of cmDN A photomodification in the induction of the suppression of skin allograft rejection in BALB/c mice preimmunized with 8-MOP/UV-A photodamaged alloreactive cells which mediates this allograft rejection. This suppression is demonstrated by inhibition of delayed-type hypersensitivity (DTH) and mixed leukocyte culture (MLC) responses. Splenocytes from BALB/c mice undergoing rejection of CBA/j skin graft which contained an expanded population of the effector T lymphocytes that mediate the rejection were treated with DNAse to remove cmDNA before or after treatment with 8-MOP and UV-A prior to infusion into naive BALB/c recipients. Mice that received pretreated effector cells were tested for MLC responses to CBA/j or B10 alloantigens before and after the DTH response. The DTH response of all groups of pretreated BALB/c mice to the relevant alloantigen was specifically suppressed as compared with the response of control mice. However, adoptive transfer of the suppression of the DTH response was optimally demonstrable only in syngeneic recipients of cells from donor mice treated with photodamaged alloreactive cells. Also, splenocytes from BALB/c mice immunized with photodamaged alloreactive cells demonstrated highly significant hyporesponsiveness and suppression of the MLC response of naive mice to the relevant alloantigen in the case of the primary MLC response, and to both alloantigens in the secondary MLC response which was totally eliminated by prior pretreatment of these effector cells with DNAse. Therefore, it appears that the suppression of the DTH response can be induced by pretreatment of the effector cells with DNAse and/or 8-MOP and UV-A but is adoptively transferable optimally only from mice which are recipients of photodamaged alloreactive cells. Moreover, the effectiveness of this treatment is decreased by prior removal of cmDNA from these cells. The presence of cmDNA is necessary for induction of suppression of the primary and secondary MLC responses in mice treated with photodamaged cells of allograft rejection.     

DNA ASSOCIATED WITH THE CELL MEMBRANE IS INVOLVED IN THE INHIBITION OF THE SKIN REJECTION RESPONSE INDUCED BY INFUSIONS OF PHOTODAMAGED ALLOREACTIVE CELLS THAT MEDIATE REJECTION OF SKIN ALLOGRAFT

Cell membrane DNA (cmDNA) is a form of DNA located on the surface of human and murine T-cells. It has recently been characterized as a target for photomodification by 8-methoxypsoralen (8-MOP) and long-wave ultraviolet light (UV-A). Whereas 8-MOP itself is biologically inert, photoactivated 8-MOP is covalently bound to pyrimidine bases in DNA. We have investigated the possible involvement of cmDNA photomodification in the induction of the suppression of skin allograft rejection in BALB/c mice preimmunized with 8-MOP/UV-A photodamaged alloreactive cells which mediates this allograft rejection. This suppression is demonstrated by inhibition of delayed-type hypersensitivity (DTH) and mixed leukocyte culture (MLC) responses. Splenocytes from BALB/c mice undergoing rejection of CBA/j skin graft which contained an expanded population of the effector T lymphocytes that mediate the rejection were treated with DNAse to remove cmDNA before or after treatment with 8-MOP and UV-A prior to infusion into naive BALB/c recipients. Mice that received pretreated effector cells were tested for MLC responses to CBA/j or B10 alloantigens before and after the DTH response. The DTH response of all groups of pretreated BALB/c mice to the relevant alloantigen was specifically suppressed as compared with the response of control mice. However, adoptive transfer of the suppression of the DTH response was optimally demonstrable only in syngeneic recipients of cells from donor mice treated with photodamaged alloreactive cells. Also, splenocytes from BALB/c mice immunized with photodamaged alloreactive cells demonstrated highly significant hyporesponsiveness and suppression of the MLC response of naive mice to the relevant alloantigen in the case of the primary MLC response, and to both alloantigens in the secondary MLC response which was totally eliminated by prior pretreatment of these effector cells with DNAse. Therefore, it appears that the suppression of the DTH response can be induced by pretreatment of the effector cells with DNAse and/or 8-MOP and UV-A but is adoptively transferable optimally only from mice which are recipients of photodamaged alloreactive cells. Moreover, the effectiveness of this treatment is decreased by prior removal of cmDNA from these cells. The presence of cmDNA is necessary for induction of suppression of the primary and secondary MLC responses in mice treated with photodamaged cells of allograft rejection.     

Silymarin, a flavonoid from milk thistle (Silybum marianum L.), inhibits UV-induced oxidative stress through targeting infiltrating CD11b+ cells in mouse skin

Phytochemicals have shown promise in inhibiting UV-induced oxidative stress, and therefore are considered as potent inhibitors of UV-induced oxidative stress-mediated skin diseases. We have shown previously that topical treatment of silymarin, a flavonoid from milk thistle (Silybum marianum), inhibits UV-induced oxidative stress in mouse skin. However, the cellular targets responsible for the inhibition of UV-induced oxidative stress by silymarin are not clearly defined. To address this issue, C3H/HeN mice were UV irradiated (90 mJ cm(-2)) with or without topical treatment with silymarin (1 mg cm(-2) skin area). Mice were killed 48 h later and skin samples collected. Flow cytometric analysis of viable dermal cells revealed that the number of infiltrating CD11b+ cells were the major source of oxidative stress (31.8%) in UV-irradiated skin compared with non-UV-exposed skin (0.4%). Treatment of silymarin inhibited UV-induced oxidative stress through inhibition of infiltrating CD11b+ cells. The analysis of myeloperoxidase also indicated that silymarin significantly (P < 0.001) decreased UV-induced infiltration of leukocytes, and this effect of silymarin was similar to that of intraperitoneal treatment of mice with monoclonal antibodies to CD11b. The inhibitory effect of silymarin, regardless of whether it is topically treated before or after UV irradiation, was of similar magnitude. Intraperitoneal administration of monoclonal antibodies to CD11b (rat IgG2b) to C3H/HeN mice inhibited UVB-induced oxidative stress generated by both epidermal and dermal cells as is evident by relative fluorescence intensity of oxidized rhodamine. Similar to the effect of anti-CD11b, silymarin also inhibited UV-induced oxidative stress in both epidermal and dermal cells. Further, CD11b+ and CD11b- cell subsets from UV-treated or silymarin+UV-treated mice were separated by immunomagnetic cell isolation technique from total epidermal and dermal single cell suspensions and analyzed for reactive oxygen species (ROS)/H2O2 production. Analytic data revealed that CD11b+ cell population from UV-irradiated skin resulted in significantly higher production of ROS in both epidermis and dermis than CD11b- cell population, and that silymarin inhibited UV-induced oxidative stress through targeting infiltrating the CD11b+ cell type in the skin.     

Solar-simulated Ultraviolet Irradiation Induces Selective Influx of CD4+ T Lymphocytes in Normal Human Skin

Abstract— The proportion and composition of the human cutaneous CD3+ T lymphocyte population was determined in situ following a single exposure to physiological, erythema-inducing doses of simulated solar radiation, mainly consisting of UV radiation. Biopsies were taken 1, 2 and 7 days after local irradiation of normal volunteers with 1,2 and 4 MED by a xenonarc lamp and immunohistochemistry was performed on cryostat sections. Ultraviolet radiation caused an initial decrease of intraepidermal CD3+ T-cell numbers or even could lead to T-cell depletion 24 and 48 h postirradiation, and this was followed by an infiltration of T cells in the epidermis as determined 1 week after UV exposure. The number of dermal CD3+ T ceDs was increased 24 h after irradiation, reached a maximum at 48 h and subsequently declined at day 7, though remained significantly higher than the unirradiated control Double staining demonstrated that the CD3+ T cells, which immigrated into the (epi)dermis upon UV exposure, coexpressed CD4 but not CD8. Therefore the CD4/CD8 ratio in skin was markedly increased during the first week upon UV exposure. Our time course study shows that UV radiation affects die T-cell population within human skin by depleting the majority of epidermal T cells and initiating a selective influx of CD4+ T cells.     

Cudraxanthone D Ameliorates Psoriasis-like Skin Inflammation in an Imiquimod-Induced Mouse Model via Inhibiting the Inflammatory Signaling Pathways

Psoriasis is a chronic inflammatory skin disease accompanied by excessive keratinocyte proliferation. Corticosteroids, vitamin D3 analogs, and calcineurin inhibitors, which are used to treat psoriasis, have diverse adverse effects, whereas natural products are popular due to their high efficiency and relatively low toxicity. The roots of the Cudrania tricuspidata (C. tricuspidata) are known to have diverse pharmacological effects, among which the anti-inflammatory effect is reported as a potential therapeutic agent in skin cells. Nevertheless, its effectiveness against skin diseases, especially psoriasis, is not fully elucidated. Here, we investigated the effect of cudraxanthone D (CD), extracted from the roots the C. tricuspidata Bureau, on psoriasis using an imiquimod (IMQ)-induced mouse model and the tumor necrosis factor (TNF)-α/interferon (IFN)-γ-activated keratinocytes. IMQ was topically applied to the back skin of C57BL/6 mice for seven consecutive days, and the mice were orally administered with CD. This resulted in reduced psoriatic characteristics, such as the skin thickness and Psoriasis Area Severity Index score, and the infiltration of neutrophils in IMQ-induced skin. CD inhibited the serum levels of TNF-α, immunoglobulin G2a, and myeloperoxidase, and the expression of Th1/Th17 cells in splenocytes. In TNF-α/IFN-γ-activated keratinocytes, CD reduced the expressions of CCL17, IL-1β, IL-6, and IL-8 by inhibiting the phosphorylation of STAT1 and the nuclear translocation of NF-kB. Taken together, these results suggest that CD could be a potential drug candidate for the treatment of psoriasis.     

Immunosuppression of xenograft rejection in porcine kidney PK15 cells by porcine IL-18

Xenotransplantation, the transplantation of cells, tissues or organs between individuals of different species, would resolve the current shortage of organs, but rejection remains the major hurdle to successful xenotransplantation. In the present study, we analyzed mixed lymphocyte reactions (MLRs) and used 51Cr release assays in order to identify the proliferation and expansion of mouse CD8+ cytotoxic T lymphocyte cells against PK15, PK15/pIL-18 or PK15/mIL-18 cells. In addition, we identified T cell populations in mouse splenocytes and lymph node cells using two-color flow cytometry. It was found that the CD8+ T cells of xenograft recipients proliferated extensively and that the survival rates of populations of PK15/mIL-18 or PK15/pIL-18 cells were higher than untransfected controls. Moreover, CD3+ T cells were increased in mice injected with PK15 cells or PK15/pIL-18 cells but PK15/pIL-18 cell numbers were lower in lymph nodes than untransfected controls. CD8+ T cells numbers were reduced in the lymph nodes of PK15/pIL-18 injected mice. These results suggest that porcine IL-18 regulates anti-pig cellular rejection in C57BL/6 mice.     

CD11b+ cells are the major source of oxidative stress in UV radiation-irradiated skin: possible role in photoaging and photocarcinogenesis

Exposure of skin to solar UV radiation induces oxidative stress and suppression of cell-mediated immune responses. These effects are associated with the greater risk of several skin disorders including photoaging and photocarcinogenesis. We have shown that UV-induced infiltrating leukocytes contribute in developing oxidative stress in UV-irradiated skin. The peak period of UV-induced infiltrating leukocytes lies between 48 and 72 h after UV exposure of the skin. In this study we demonstrated that UV (90 mJ/cm2)-induced infiltrating CD11b+ cells in C3H/HeN mice skin were the major source of oxidative stress. Hydrogen peroxide (H2O2) was determined as a marker of oxidative stress. Flow cytometric analysis of viable cells revealed that the number of CD11b+H2O2+ cells were significantly higher (31.8%, P < 0.001) in UV-irradiated skin in comparison with non-UV-exposed skin (0.4%). Intraperitoneal administration of monoclonal antibodies to CD11b (rat IgG2b) to C3H/HeN mice inhibited UVB-induced infiltration of leukocytes, as evidenced by reduction in myeloperoxidase activity (64-80%, P < 0.0005), concomitant with significant reduction in H2O2 production both in epidermis and dermis (66-83%, P < 0.001-0.0005) when compared with the administration of rat IgG2b isotype of anti-CD11b. Furthermore, CD11b+ and CD11b- cell subsets were separated by immunomagnetic cell isolation technique from total epidermal and dermal single cell suspensions obtained 48 h after UV irradiation of the skin and analyzed for H2O2 production. Analytical data revealed that CD11b+ cell population from UV-irradiated skin resulted in significantly higher production of total H2O2 in both epidermis and dermis (87-89%, P < 0.0001) in comparison with CD11b- cell population (11-13% of total H2O2). These data revealed that infiltrating CD11b+ cells were the major source of oxidative stress in UV-irradiated skin and thus may contribute to photoaging and promotion of skin tumor growth within the UV-irradiated skin. Together, these data suggest that reduction in UV-induced skin infiltration of CD11b+ cells may be an alternative and effective strategy to reduce solar UV light-induced oxidative stress-mediated skin disorders including photoaging and photocarcinogenesis.     

IL-17-deficient allogeneic bone marrow transplantation prevents the induction of collagen-induced arthritis in DBA/1J mice

IL-17-producing CD4⁺ T cells (Th17) play important functions in autoimmune diseases and allograft rejection of solid organs. We examined the effects of IL 17 and its mechanism of action on arthritis in a murine collagen-induced arthritis (CIA) model using bone marrow transplantation (BMT) system. DBA/1J mice were administered a lethal radiation dose and then rescued with bone marrow derived from either wild-type (WT) or IL-17^-/- mice on C57BL/6 background mice. CIA was induced after the bone marrow transplant, and disease progression was characterized. DBA/1J mice with CIA that received IL-17^-/- donor bone marrow showed potently inhibited development and severity of clinical arthritis as compared with CIA mice that received WT bone marrow. Reduced secretion of the pro-inflammatory cytokines tumor necrosis factor-α, IL-1β, and IL-6, and collagen-specific T cell responses were observed in mice that received IL-17^-/- bone marrow. IL-17 blockade also inhibited effector T cell proliferation by reciprocally regulating the Treg/Th17 ratio. IL-17 blockade prevented joint destruction in mice with CIA. These findings suggest that CIA with BMT is a viable method of immunological manipulation and that IL-17 deficiency suppresses severe joint destruction and inflammation in CIA mice. There may be clinical benefits in blocking IL-17 and BMT in the treatment of rheumatoid arthritis.     

Higher infiltration by Th17 cells compared with regulatory T cells is associated with severe acute T-cell-mediated graft rejection

The aim of this study was to evaluate whether the Th17 and Treg cell infiltration into allograft tissue is associated with the severity of allograft dysfunction and tissue injury in acute T cell-mediated rejection (ATCMR). Seventy-one allograft tissues with biopsy-proven ATCMR were included. The biopsy specimens were immunostained for FOXP3 and IL-17. The allograft function was assessed at biopsy by measuring serum creatinine (Scr) concentration, and by applying the modified diet in renal disease (MDRD) formula, which provides the estimated glomerular filtration rate (eGFR). The severity of allograft tissue injury was assessed by calculating tissue injury scores using the Banff classification. The average numbers of infiltrating Treg and Th17 cells were 11.6 ± 12.2 cells/mm2 and 5.6 ± 8.0 cells/mm2, respectively. The average Treg/Th17 ratio was 5.6 ± 8.2. The Treg/Th17 ratio was significantly associated with allograft function (Scr and MDRD eGFR) and with the severity of interstitial injury and tubular injury (P < 0.05, all parameters). In separate analyses of the number of infiltrating Treg and Th17 cells, Th17 cell infiltration was significantly associated with allograft function and the severity of tissue injury. By contrast, Treg cell infiltration was not significantly associated with allograft dysfunction or the severity of tissue injury. The results of this study show that higher infiltration of Th17 cell compared with Treg cell is significantly associated with the severity of allograft dysfunction and tissue injury.     

Noninvasive Early Diagnosis of Allograft Rejection by a Granzyme B Protease Responsive NIR-II Bioimaging Nanosensor

Early diagnosis of allograft rejection helps to improve the immune-related management of transplant recipients. The clinically-used core needle biopsy method is invasive and subject to sampling error. In vivo fluorescence imaging for monitoring immune-related processes has the advantages of non-invasiveness, fast feedback and high sensitivity. Herein, we report a responsive second near-infrared (NIR-II) fluorescent nanosensor (ErGZ) to detect early allograft rejection. ErGZ allows ratiometric in vivo fluorescence sensing of granzyme B, which is overexpressed in recipients’ T cells during the onset of rejection. The sensor demonstrates efficacious detection of allograft rejection with high sensitivity and specificity, which accomplishes non-invasive diagnosis of rejection in skin and deep buried islets transplant mice models 2 d and 5 d earlier than biopsy, by in vivo fluorescence imaging and urinary detection, respectively, providing a valuable approach for therapeutical management.     

Ultraviolet light induces reactivation in a murine model of cutaneous herpes simplex virus-1 infection.

We have developed a model of cutaneous herpes simplex virus-1 (HSV-1) reactivation in SKH-1 hairless mice which closely mimics the condition in humans. Sixty plaque-forming units of HSV-1 strain 17 syn+ were applied to a superficially abraded area on the lateral body wall. More than 85% of mice developed primary HSV-1 infection characterized by a zosteriform pattern of cutaneous vesiculation and ulceration. Approximately one-third of mice with primary skin lesions succumbed to neurologic disease and in the remaining mice cutaneous lesions healed completely. Subsequent exposure of healed areas to two minimal inflammatory doses of UV resulted in recrudescence of skin lesions in the irradiated areas in almost 60% of mice. Lesions appeared approximately 4 days after irradiation, persisted for 3-5 days and then resolved completely. Reactivation rarely resulted in death due to neurologic disease. Primary lesions had a histologic appearance typical of cutaneous HSV-1 infection with vesicles and focal epithelial necrosis accompanied by the formation of epithelial syncytial cells and the presence of herpetic intranuclear inclusion bodies. In primary lesions HSV-1 was demonstrated by immunohistochemistry, polymerase chain reaction and culture. In reactivated lesions epithelial syncytia and inclusion bodies were not seen; however, virus was demonstrable by polymerase chain reaction and culture. Exposure of the uninfected side to UV did not stimulate disease recurrence suggesting that local effects of UV rather than systemic immunosuppression were responsible for reactivation. Reactivation could also be obtained with two minimal inflammatory doses of UV from a UV-340 light source which emits light approximating the solar spectrum.     

IL-10 does not play a role in cutaneous Photofrin photodynamic therapy-induced suppression of the contact hypersensitivity response.

Photodynamic therapy (PDT) treatment of both malignant and benign skin diseases has proven to be effective, and its use is increasing worldwide. However, preclinical studies using murine models have shown that PDT of the skin inhibits cell-mediated immune reactions, as measured by the suppression of the contact hypersensitivity (CHS) reaction. We have previously demonstrated that PDT enhances IL-10 expression in treated skin, and that the kinetics of induction of IL-10 is similar to the kinetics of suppression of systemic CHS reactions by cutaneous PDT. In the following report we have expanded upon these studies to demonstrate that cutaneous PDT, using Photofrin, induces elevated levels of systemic IL-10 that persist for at least 28 days following treatment. The increase in systemic IL-10 correlates to a prolonged suppression of CHS of at least 28 days following cutaneous PDT. IL-10 has been implicated as the causative agent in the suppression of cell-mediated immune reactions by UVB and transdermal PDT. However, in the studies reported here we demonstrate that the suppression of CHS by cutaneous PDT occurs via an IL-10 independent mechanism, as administration of anti-IL-10 antibodies had no effect on the ability of PDT to induce CHS suppression. These results were further confirmed using IL-10 knockout (KO) mice. Cutaneous PDT of IL-10 KO mice resulted in CHS suppression that was not significantly different from suppression induced in wild-type mice. Thus, it appears as though IL-10 does not play a role in CHS suppression by cutaneous PDT. Suppression of cell-mediated immune reactions by UVB and transdermal PDT is reversible by IL-12, which is critical for the development of these reactions. We show that administration of exogenous IL-12 is also able to reverse CHS suppression induced by cutaneous PDT, suggesting that whereas suppression of cell-mediated immune reactions by UVB, transdermal PDT and cutaneous PDT occurs via different mechanisms, a common regulatory point exists.