Experimental validation of the filtering approach for dose monitoring in proton therapy at low energy.

The higher physical selectivity of proton therapy demands higher accuracy in monitoring of the delivered dose, especially when the target volume is located next to critical organs and a fractionated therapy is applied. A method to verify a treatment plan and to ensure the high quality of the hadrontherapy is to use Positron Emission Tomography (PET), which takes advantage of the nuclear reactions between protons and nuclei in the tissue during irradiation producing beta(+)-emitting isotopes. Unfortunately, the PET image is not directly proportional to the delivered radiation dose distribution; this is the reason why, at the present time, the verification of depth dose profiles with PET techniques is limited to a comparison between the measured activity and the one predicted for the planned treatment by a Monte Carlo model. In this paper we test the feasibility of a different scheme, which permits to reconstruct the expected PET signal from the planned radiation dose distribution along beam direction in a simpler and more direct way. The considered filter model, based on the description of the PET image as a convolution of the dose distribution with a filter function, has already demonstrated its potential applicability to beam energies above 70 MeV. Our experimental investigation provides support to the possibility of extending the same approach to the lower energy range ([40, 70] MeV), in the perspective of its clinical application in eye proton therapy.

Two-dimensional dosimetry of radiotherapeutical proton beams using thermoluminescence foils.

In modern radiation therapy such as intensity modulated radiation therapy or proton therapy, one is able to cover the target volume with improved dose conformation and to spare surrounding tissue with help of modern measurement techniques. Novel thermoluminescence dosimetry (TLD) foils, developed from the hot-pressed mixture of LiF:Mg,Cu,P (MCP TL) powder and ethylene-tetrafluoroethylene (ETFE) copolymer, have been applied for 2-D dosimetry of radiotherapeutical proton beams at INFN Catania and IFJ Krakow. A TLD reader with 70 mm heating plate and CCD camera was used to read the 2-D emission pattern of irradiated foils. The absorbed dose profiles were evaluated, taking into account correction factors specific for TLD such as dose and energy response. TLD foils were applied for measuring of dose distributions within an eye phantom and compared with predictions obtained from the MCNPX code and Eclipse Ocular Proton Planning (Varian Medical Systems) clinical radiotherapy planning system. We demonstrate the possibility of measuring 2-D dose distributions with point resolution of about 0.5 x 0.5 mm(2).

Intracerebral recruitment and maturation of dendritic cells in the onset and progression of experimental autoimmune encephalomyelitis.

Dendritic cells (DCs) are thought to be key elements in the initiation and maintenance of autoimmune diseases. In this study, we sought evidence that DCs recruited to the central nervous system (CNS), a site that is primarily devoid of resident DCs, play a role in the effector phase and propagation of the immune response in experimental autoimmune encephalomyelitis (EAE). After immunization of SJL mice with proteolipid protein 139-151 peptide, process-bearing cells expressing the DC markers DEC-205 and CD11c appeared early in the spinal cord. During acute, chronic, and relapsing EAE, DEC-205(+) DCs expressing a lymphostimulatory phenotype (including the mature DC marker MIDC-8, major histocompatibility complex class II, CD40, and CD86 molecules) accumulated within the CNS inflammatory cell infiltrates. More prominent infiltration of the spinal cord parenchyma by mature DCs was observed in mice with relapsing disease. Macrophage inflammatory protein 3alpha, a chemokine active on DCs and lymphocytes, and its receptor CCR6 were up-regulated in the CNS during EAE. These findings suggest that intracerebral recruitment and maturation of DCs may be crucial in the local stimulation and maintenance of autoreactive immune responses, and that therapeutic strategies aimed at manipulating DC migration could be useful in the treatment of CNS autoimmune disorders.

Persisting viruses and autoimmunity.

Viral infections can be responsible for the onset and sustaining of autoimmune processes. We discuss how chronic inflammation associated with viral persistence is the prerequisite for initiation of a multi-step process leading to autoimmunity. Firstly, chronic inflammation may favor the priming of autoreactive T cells that have escaped thymic selection and are specific for self-mimicking viral peptides in the periphery. In addition, viral persistence and inflammation can act synergistically to induce and sustain autoimmunity either unveiling cryptic self-epitopes, or favoring determinant spreading, or activating dendritic cells, or promoting constant priming of new autoreactive T cells, or contributing to the efficient generation of effector cells, or, finally, restimulating memory T lymphocytes.

Short-lived immunization site inflammation in self-limited active experimental allergic encephalomyelitis.

To understand the mechanisms underlying spontaneous remission of proteolipid protein (PLP) 139-151 peptide-induced experimental allergic encephalomyelitis (EAE), an acute autoimmune disease of SJL mice resembling human multiple sclerosis, we examined both the effector response site in the central nervous system (CNS) and the immunization site at different phases of the disease. In the CNS, the frequency of PLP 139-151 peptide-specific IFN-gamma-producing T cells as well as the amount of infiltrating CD4(+) and CD11b(+) cells decreased with recovery. However, IL-4-producing cells were always rare and cyclooxygenase-2(+) cells were numerous only at disease peak in the CNS, suggesting that T(h)2 cytokines and prostaglandins did not determine remission of EAE. By looking at the s.c. site of PLP 139-151 peptide plus adjuvant injection, we found that, although the inflammatory infiltrate was abundant, CD11b(+) cells started to decrease already during disease acute phase and DEC-205(+) cells were numerous only at early time points. We propose that immunization site inflammation is short-lived in PLP 139-151 peptide-induced EAE, and this leads to a temporary autoreactive T cell stimulation and to a self-limited disease.

On the lifespan of virgin T lymphocytes.

To study the lifespan of virgin T lymphocytes, we removed the thymus from adult female mice and then, at various times afterward, tested their ability to mount an immune response to a newly encountered Ag, the male Ag H-Y. We found that unprimed thymectomized mice were able to generate a primary response to H-Y for some time after thymectomy but lost this ability at approximately 6 mo. In contrast, mice that were primed to H-Y just after thymectomy continued to display immunological memory to H-Y for >1 year. These experiments show that primary immune responses disappear in the absence of a thymus.

Lack of Th2 cytokine increase during spontaneous remission of experimental allergic encephalomyelitis.

The mechanisms underlying spontaneous remission of autoimmune diseases are presently unknown, though regulatory T cells are believed to play a major role in this process. We tested the hypothesis that Th2 and/or other T cell regulatory cytokines cause the spontaneous remission of experimental allergic encephalomyelitis (EAE), a model of Th1-mediated autoimmunity. We analyzed the cytokine profile of lymph node and central nervous system-infiltrating cells in individual SJL mice at different stages of proteolipid protein (PLP) 139-151 peptide-induced EAE. We found that IFN-gamma slowly fades away after clinical recovery, whereas IL-4, IL-10 and transforming growth factor-beta remain low or undetectable. Our peptide-results therefore suggest that regulatory T cells producing anti-inflammatory cytokines are not involved in spontaneous remission of EAE and challenge the view that the Th1/Th2 balance has a key role in EAE regulation.

Persisting viruses and chronic inflammation: understanding their relation to autoimmunity.

Viral infections may induce and sustain autoimmune processes via several and overlapping mechanisms. We outline how chronic inflammation, sustained by persisting viruses, may be "the prerequisite" for initiation and maintenance of the multistep process leading to autoimmunity. Chronic inflammation may favour priming of autoreactive T cells which have escaped thymic tolerance and are able to mount a cross-reactive response to self-mimicking antigens carried by viruses in the periphery. Moreover, chronic inflammation and persisting viruses can synergistically support autoimmunity through other relevant mechanisms: unveiling of cryptic self-epitopes, determinant spreading, activation of dendritic cells, constant priming of new autoreactive T cells, and efficient generation and restimulation of memory cells. Therefore, viruses seem to play a key role among the many environmental factors which, together with the genetic background, have been implicated in the pathogenesis of autoimmune diseases. We will also discuss some hypotheses explaining why autoimmunity is a rare event.

A conditioned dendritic cell can be a temporal bridge between a CD4+ T-helper and a T-killer cell.

To generate an immune response, antigen-specific T-helper and T-killer cells must find each other and, because they cannot detect each other's presence, they are brought together by an antigen-loaded dendritic cell that displays antigens to both. This three-cell interaction, however, seems nearly impossible because all three cell types are rare and migratory. Here we provide a potential solution to this conundrum. We found that the three cells need not meet simultaneously but that the helper cell can first engage and 'condition' the dendritic cell, which then becomes empowered to stimulate a killer cell. The first step (help) can be bypassed by modulation of the surface molecule CD40, or by viral infection of dendritic cells. These results may explain the long-standing paradoxical observation that responses to some viruses are helper-independent, and they evoke the possibility that dendritic cells may take on different functions in response to different conditioning signals.

Long-lasting CD8 T cell memory in the absence of CD4 T cells or B cells.

The cellular basis of immunological memory has been a debated issue. It is not clear whether CD8 T cell memory is maintained by long-lived cells or by specific or nonspecific restimulation. Here, we have approached the question from a different angle, asking whether the cellular interactions that are required to maintain memory are the same as those necessary to activate cytotoxic T lymphocytes. We studied the CD8 memory response to the male antigen H-Y in mice deficient in CD4 cells, or B cells and found that memory in these mice was virtually unimpaired. These results suggest that CD8 memory is CD4 independent and that there is no requirement for long term retention of immune complexes on follicular dendritic cells, nor for B cells as antigen-presenting cells.

Successful T cell priming in B cell-deficient mice.

B cells are an abundant population of lymphocytes that can efficiently capture, process, and present antigen for recognition by activated or memory T cells. Controversial experiments and arguments exist, however, as to whether B cells are or should be involved in the priming of virgin T cells in vivo. Using B cell-deficient mice, we have studied the role of B cells as antigen-presenting cells in a wide variety of tests, including assays of T cell proliferation and cytokine production in responses to protein antigens, T cell killing to minor and major histocompatibility antigens, skin graft rejection, and the in vitro and in vivo responses to shistosome eggs. We found that B cells are not critical for either CD4 or CD8 T cell priming in any of these systems. This finding lends support to the notion that the priming of T cells is reserved for specialized cells such as dendritic cells and that antigen presentation by B cells serves distinct immunological functions.

HLA class II molecules transduce accessory signals affecting the CD3 but not the interleukin-2 activation pathway in T blasts.

MHC class II molecules play a central role in the control of the immune response, but their biologic function and mechanism of action on the surface of activated human T lymphocytes are not entirely understood. In our study, the functional role of HLA class II molecules in T-blast proliferation was investigated by analyzing in parallel the IL-2- and CD3-driven activation pathways. The results indicate that the cross-linking of class II and CD3 molecules significantly increased the CD3-mediated T-blast proliferation, while no effect was observed on the IL-2-driven cell activation. This phenomenon was not confined to either CD4+ or CD8+ subsets nor was specifically affected by CD45 triggering. Biochemical studies showed that signaling via MHC class II molecules in T blasts led to PKC membrane translocation and IP accumulation. The simultaneous triggering of CD3 and HLA class II molecules led to a synergistic effect on IP accumulation but did not increase the CD3-mediated PKC membrane translocation. Our data suggest that HLA class II molecules are involved in T-cell-T-cell interactions and can mediate accessory signals, affecting the T-lymphocyte activation state.

Free microvascular fibular versus conventional bone grafts.

Free microvascular grafts have many advantages over conventional bone grafts. Above all they do not undergo creeping substitution and live on their own vessels, thus being able to heal quickly and fight infection. Research in rabbits has been done with radiographic, scintigraphic and light and fluorescence histologic examination which demonstrated the high superiority of quality, reliability and rapidity in healing of the microvascular bone transfer as related to avascular ones. Eighty-five cases of free microvascular fibular transfers starting in 1975 are presented including six congenital cases, 12 non-unions, eight osteitis, seven tumors, seven large losses of bone and 45 femoral head necrosis. The failure ratio is very low. Six primary failures were cured by plaster or additional cancellous bone grafts. Four failures (in femoral head necrosis) did not cure due to recession of the graft, its reabsorption or progression of avascular necrosis in two corticosteroid cases.

Neurolized nerve padding in actinic lesions: omentum versus muscle use. An experimental study.

Free microvascular greater omentum flap to pad brachial plexus after neurolysis for actinic lesions is a major operation. Thus several authors have suggested a simple padding by means of traditional muscle flaps. As contouring the brachial plexus elements is difficult using muscle, we wished to evaluate the formation of perineural fibrosis with either omentum or muscle, following neurolysis. An experimental model in rats was set up, dividing the two femoral nerves and wrapping one of them with the greater omentum and the other with abdominal muscles. The degree of perineural fibrosis at light microscopy observation was fivefold greater when muscle, rather than omentum, was used.