The Expression Of Id2 In Normal Adult And EAE Rat Spinal Cord And Progesterone Treatment Changes

Multiple sclerosis (MS) is a central nervous system (CNS) demyelinating disease with teenage incidence, which is characterized by multiple lesions; the incidence can be as high as 0.05-0.1%. MS mostly affects people aged from 20 to 40 with a peak at 30. Most of the patients left sequels, with the increased frequency of seizures, sequels in a steadily increasing. Typical cases are remission– relapse and stepped deterioration. The mechanism by which MS occurs is unknown, and the typical pathological changes mainly include the spinal cord, brain stem, cerebellum, and cerebral white matter, visible multiple demyelinating lesions. Recent evidence has shown that oligodendrocyte precursor cells (Opcs) and immature oligodendrocytes (OL) are preserved in MS lesions, playing roles in pathogenesis of MS. Induced differentiation of these cells into myelinating cells may ultimately lead to a novel remyelination therapy of MS. Though a wide variety of therapeutic strategies have already been proposed and adopted for MS, none of those available drugs are specific. For example, adrenocorticotropic hormone and corticosteroid were proved helpful in shortening the acute episode, but no effect was show in the whole pathogenesis and the long-term prognosis. Immunosuppressive agents that can be used, but have no effects on the whole course of MS. Other therapies based on immunoglobulins, recombinant interferon (IFN), plasma exchange, antilymphocyte serum, etc have been proved to be effective in a subset of patients with MS, however, there are many problems to be tacked such as side effects, safety, high cost of prolonged treatment and methodological limitations, so it is still essential to find novel therapeutic strategies for MS.Studies find that the ethidium bromide injection induces demyelinating CNS model with the intervention of progesterone the ratio of demyelination axons decreased and OL involved in the regeneration of myelin increased significantly. Progesterone significantly increases an older female rat cerebellar peduncle remyelination of axons. It can reduce the neurological symptoms of brain injury and accelerate the formation of in vitro model of cerebellar myelin. No one known whether progesterone can be a targeted therapeutic drugs or not. Although the mechanism is still unknown, in 2005 Scotland has begun a large-scale clinical research, giving postpartum patients'large doses of progesterone treatment, Observing the relapse prevention and effects of progesterone. This research is expected to be finished in 2008.DNA-binding inhibitor (Id) is the transcriptional inhibitors of basic helix - loop-helix (bHLH) containing HLH domain proteins. As a factor which inhibit bHLH protein lacks the DNA binding basic sequenceId2 is generally synthesized by myogenic precursor cells. Id2 regulates the development and differentiation in diverse cell types. Id2 functions as the blocks of cell differentiation and is reduced during the course of cell differentiation. Id2 participates in the development of T cells, tumors cell proliferation, cell cycle and muscle genesis and so on. In recent years, many evidences provide that Id2 plays an important role in nervous system. In Id2 null mice, there are many abnormities such as immature neuralblast. It is reported that Id2 is expressed in ependymal cell, dorsal root ganglion sensory neural progenitor in peripheral nervous system by in situ hybridization. All these evidences show that Id2 plays an important role in nervous system. After spinal cord injury around the central canal Nestin-positive cells also express Id2 mRNA. This suggests that Id2 is also express in neural progenitor cells, which progenitor cells have the potential to differentiate into many cells, suggesting that Id2 may play an important role in the repair of nerve system. Researches have shown that progesterone can promote OL cell line development, and the mature OL related gene expression. Thus enabling the regeneration of myelin speed. At the differentiation onset of oligodendrocyte precursor cells to oligodendrocyte Id2 transfer from the cytoplasm to the nucleus. Over-Expression Id2 almost completely block the differentiation of oligodendrocyte precursor cells into oligodendrocytes, the Id2-/- Opcs can not differentiate into oligodendrocytes maturely. Id2 plays an important role in the oligodendrocyte differentiation. The experiment is purposed to find out whether the progesterone mechanism can affect oligodendrocyte differentiation by regulating the expression of Id2.Experimental autoimmune encephalomyelitis (EAE) is an effective model for studying multiple sclerosis (MS), therefore, we used fluorescence and immunohistochemical technique to examine the expression of Id2 in normal and the different time of EAE rat spinal cord. In addition the expression of Id2 by progesterone treatment in EAE different time with the results as follows:1. The expression of Id2 in normal rat spinal cordId2 positive cells are extensively distributed in normal adult rat of spinal cord in cervical segments, thoracic segments, lumbar segments and sacral segments of the gray and white matter .Both cytoplasm and nucleus are seen.1.1The gray matter: there is no difference among the segmental distributions. In laminaⅠ-Ⅲthe expression of Id2 is predominantly in nucleus but few in cytoplasm. The numbers and staining of positive cells increases from laminaⅠto laminaⅢgradually. Cytoplasmic staining positive cells of a smaller volume appear in laminaⅣ. The expression of Id2 also can be seen in nucleus. Cytoplasmic staining with a monoploar and bipolar morphology cells appears in laminaⅤ-Ⅶ, the numbers and the volume of staining of positive cells noticeably increases in laminaⅤand laminaⅦ. In laminaⅦalso can be seen with beaded positive cells, the expression of Id2 also can be seen in nucleus. The morphology in laminaⅧis similar to laminaⅦ, but the cytoplasmic staining positive cell has a larger volume than laminaⅦ, dyeing deeper than laminaⅦ, the expression of Id2 also can be seen in nucleus. In laminaⅨthe expression of Id2 was predominantly in nucleus but few in cytoplasm .In intermediomedial cell column, (IMM) and intermediolateral cell column (IML) the expression of Id2 both cytoplasm and nucleus.1.2 The white matter: the expression of Id2 spread widely in white matter glial cells and nerve fibers, and the staining of positive cells deeper in lateral spinal nucleus (Lsp) and the pyramidal tract (PY) is visible.1.3 fluorescence Results: GFAP+ astrocytes in the spinal cord are widely distributed. High-power microscope extends long neurite. The distribution of CC-1+ oligodendrocyte in the spinal cords extensively, mainly in the cytoplasm, the cell bodies of small, protruding short.Id2 co- expression with CC-1 +oligodendrocyte and do not express in GFAP+ astrocytes andβ-tubulin+ neuron.2. The expression of Id2 in EAE rat of spinal cord at different time points the expression of Id2 extensively distributed in EAE rat of spinal cord in cervical segments, thoracic segments, lumbar segments and sacral segments of the gray and white matter, Mainly expressing in the nucleus but few in cytoplasm. Comparing the expression of Id2 in EAE model at different time points to the normal control group and adjuvant control group, were significantly different (P <0.01) there is no significant difference between normal control group and adjuvant group (P> 0.05). IOD value lower expression in normal rat spinal cord. The expression of EAE model at different time points IOD value is higher than those of the normal group and adjuvant control group (P <0.01). In six days the expression of come to its maximum, and then it decreases gradually. The lowest expression appears after eight days. then increased gradually, in 10th,15th still see higher level of expression.That fluorescent marker shows CC-1 +oligodendrocyte also expresses Id2 indicates that the oligodendrocyte expression Id2 in EAE model.3. The expression of Id2 in Progesterone treatment group of spinal cord the expression of Id2 extensively distributed in Progesterone treatment group at different times in spinal cord cervical segments, thoracic segments, lumbar segments and sacral segments of the gray and white matter. mainly expressed in the nucleus and few in cytoplasm .By progesterone therapy six days Id2 higher expression in the spinal cord, eight days dropped significantly, 10,15 days gradually restored to the level at the beginning of treatment.Progesterone treatment Id2 expression of the statistical resultsThe expression of Id2 in EAE model is higher than the normal rats, after progesterone treatment, the expression of Id2 was significantly lower (P <0.05). In six days of treatment group the expression of Id2 decreases most (P <0.01). In 15th, the decrease compare to EAE group there still have significant difference (P <0.05).Conclusions:1. With the method of Double-labeled immunofluorescent staining combined with enzyme labeled assay the feature of Id2 expression in normal adult rat spinal cord. The results indicate that Id2 wildly distributed in spinal cord segments,different regions of gray matter and white matter .It may be involved in the process of central nerve system myelination and oligodendrocyte regeneration. 2. The temporal and spatial distributions of Id2 in rat spinal cord is in consonance with the pathogenesis of EAE characterized by a relapse-remission course and progressive deterioration, suggesting the pathogenesis of multiple sclerosis may closely relate to Id2.3. After Progesterone treatment, the expression of Id2 is significantly lower indicate that therapic mechanism of progesterone on the multiple sclerosis may affect the expression of the Id2.4. We identify Id2 was mainly expressed in oligodendrocyte suggested a potential role of Id2 in the differentiation and proliferation of oligodendrocyte which was responsible for central nerve myelination.