The Clinical And Basic Research Of Cushing’s Disease With Hyperprolactinemia

Part 1 Clinical Research of Cushing’s Disease with HyperprolactinemiaBackgroundPituitary adenoma is a common tumor of intracranial benign tumors. About 10% of intracranial tumors are pituitary adenomas. The mass effect, hormone secretion dysfunction and postoperative complications can severely impact the patient’s quality of life and even life-threatening. Therefore, the early and correct diagnosis and treatment of pituitary adenoma is very necessary and important. About 5% to 15% of functional pituitary adenomas are corticotropic pituitary adenomas. Nowadays, studies on pituitary adenomas are common, but it is rare in Cushing’s disease with hyperprolactinemia. Therefore, the characteristics of Cushing’s disease with hyperprolactinemia were not clearly established, probably due to the relatively low annual incidence. The aim of this study is to evaluate the clinical data from a large cohort of merely Cushing’s Disease, merely prolactin-secreting adenomas and Cushing’s disease with hyperprolactinemia.ObjectiveIn this part, we focus on the differences between groups in genders, age, clinical symptoms, course of disease, hormone levels, imaging performance and postoperative recurrence and etc. Statistical analysis used to explore the differences between merely Cushing’s Disease, merely prolactin-secreting adenomas and Cushing’s disease with hyperprolactinemia. We hope to improve the comprehension on Cushing’s disease with hyperprolactinemia, and expect to be able to improve the diagnosis and treatment of Cushing’s disease with hyperprolactinemia.MethodsWe selected patients treated with surgery and diagnosed to be merely Cushing’s Disease, merely prolactin-secreting adenomas and Cushing’s disease with hyperprolactinemia by pathology in Neurosurgery, Department of Shandong Provincial Hospital from 2005 to 2013. SPSS 16.0 statistical analysis software was used to analyze data in every program. Chi-square test, Fisher’s Exact test, t-test, Mann-Whitney test, Bonferroni correction, and Bivariate correlation were used to analyze data. And P<0.05 was considered as statistically significant.Results1. Compared with CD group, patients in CD+PRL group were characterized by younger age, larger tumor size, higher rates of invasion according to Knosp classification, more clinical manifestations (including a higher incidence of menstrual disorders caused by PRL secretion), lower endocrine normalization rate. However, the incidences of progressive obesity and hypertension were much lower in patients in the CD+PRL group than in patients in the CD group. Tumor recurrence was significantly less frequent in the CD group (8.3%) than in the CD+PRL group (36.1%).2. Compared to patients in the PRL groups, patients in the CD+PRL were younger, had higher incidences of headaches and dizziness, progressive obesity, menstrual disorders, polyuria and polydipsia, hypertension, diabetes mellitus, and had higher rates of postoperative hyponatremia. Recurrence rates in the CD+PRL groups were higher than those in the CD group. However, endocrine normalization rate in the CD+PRL groups was lower.Conclusions1. Patients in the CD+PRL group were characterized by younger age, larger tumor size, more clinical manifestations (including a higher incidence of menstrual disorders caused by PRL secretion), lower endocrine normalization rate and more frequent recurrence rate, compared with patients with Cushing’s disease alone.2. Patients in the CD+PRL group were younger, had more complications, had lower endocrine normalization rate and had more frequent recurrence rate compared with PRL patients.Part 2 Basic Research of Cushing’s Disease with Hyperprolactinemia Chapter 1 Cellular Immunity and Humoral Immunity of Cushing’s Disease with HyperprolactinemiaBackgroundThe neuro-endocrine-immunoregulatorynetwork (NEI) is fundmnental to the maintenance of homeostasis. Recent evidence suggests a two-way effect between the neuroendocrine system and immune system. Immune cells can secrete neurotransmitters and endocrine hormones. At the same time, it can identify the signal molecules produced by central nervous system and endocrine system through corresponding receptors. Cushing’s disease is a rare disorder characterized by overproduction of ACTH from a pituitary adenoma leading to hypercortisolemia that in turn leads to increased morbidity and mortality. Hyperprolactinemia (HPRL) can cause metabolic and inflammatory changes which are associated with accelerated atherosclerotic process. Cushing’s disease with hyperprolactinemia due to mixed ACTH-and PRL-secreting adenomas occurs rarely. Very few investigations have focused on the cellular immunity and humoral immunity of these patients.ObjectiveThis chapter investigated the changes of humoral immunity and cellular immunity in the way of evaluating the levels of IgG, IgA, lgM and C3, C4, T-lymphocyte, B-lymphocyte Subpopulations, NKcells in peripheral blood of Cushing’s disease with hyperprolactinemia. In addition, we studied the positive rate of antinuclearantibody in the serum of these patientsin order to search for the possibility of anti-nuclear antibody as tumor markers.Methods1. Subjects:Twenty patients with Cushing’s disease with hyperprolactinemia and forty patients with Cushing’s disease as controls were selected in this study.2. The conients of IgG, IgA, lgM and C3, C4 were examined by rate nephelometry. The levels of CD3+T, CD4+T, CD8+T-lymphoeytes, CD16+CD56+NK, CD19+ B-lymphoeytes were tested with the method of floweytometry.3. Used indirect immunofluorescence to detect the cases with antinuclear antibodies.4. Analyses were performed using Statistical Package for Social Sciences Version 16.0 (Chicago, IL, USA). All statistical tests were two-sided, and P<0.05 was considered statistically signiicant.Results1. No significant difference in the levels of IgG, IgA, lgM and C3, C4 was observed between Cushing’s disease patients with hyperprolactinemia and the controls (P>0.05).2. The percentage of CD3+T cells, CD4+T cells, CD3-CD16/56+NK cells decreased in Cushing’s disease patients with hyperprolactinemia compared to normal controls (P＜0.001, P＜0.001, P＜0.001). The CD4+T/CD8+T ratio was also significantly redueed(P<0.001).3. No significant difference in the rate of antinuclear antibody was observed between Cushing’s disease patients with hyperprolactinemia and the controls (P=0.439).Conclusion1. There was only a minor effect to humoral immunity.2. The patients mainly demonstrated the decrease of T-helper lymphocytes as the sign of cellular immunity function disorders.3. No significant difference in the rate of antinuclear antibody was observed between Cushing’s disease patients with hyperprolactinemia and the controls.Chapter 2 Association of killer cell immunoglobuiin-like Receptors and its HLA-C Ligand in Cushing’s Disease with HyperprolactinemiaBackgroundHormone achieved the function of the immune system mainly through the regulation of immune cells. In recent years, the onset of pituitary adenoma is obviously rising. Killer cell immunoglobulin-like receptors (KIRs) are a highly diverse family of receptors expressed by natural killer cells and subsets of T lymphocytes. KIR molecule belongs to immuneglobulin superfamily and modulates cells function upon recognition of HLA class I molecules. Previous studies have demonstrated that KIR genes are involved in the pathogenesis of a variety of diseases. However up to now, the role of KIR polymorphisms and HLA-Cw alleles in patients Cushing’s disease patients with hyperprolactinemia has not been investigated in China. Therefore, we can infer that KIR gene polymorphisms may exert a crucial role in Cushing’s disease patients with hyperprolactinemia.ObjectiveTo explore whether KIR gene and HLA-Cw alleles polymorphisms are associated with susceptibility to Cushing’s disease patients with hyperprolactinemia. And provide a better understanding on the genetic diversity of KIR across these patients.Methods1. Subjects:The 100 samples analyzed for this study comprised 20 Cushing’s disease patients with hyperprolactinemia and 80 controls.2. Genome DNA extraction:Genomic DNA sample was extracted from thylene diamine tetraacetic acid (EDTA) anticoagulated peripheral blood with a TIANamp Blood DNA kit and stored at -20℃ before use.3. KIR genotyping:Genotyping of KIR was conducted by SSP-PCR method, which was performed to detect the presence or absence of KIR genes. So KIR locus typing was performed to detect the presence or absence of 6 inhibitory KIR genes (2DLI,2DL2, 2DL3,2DL5,3DL1,3DL3),6 activating KIR genes (2DS1, DS2,2DS3,2DS4,2DS5, 3DS1). PCR cycle reactions were performed on a Gene Amp PCR System 9700.4. HLA-Cw genotyping:Genotyping of HLA-Cw was also conducted by PCR-SSP method. KIR locus typing was performed to detect the presence or absence of HLA-Cw*01-08. PCR cycle reactions were performed on a Gene Amp PCR System 9700.5. Statistical analysis:Analysis was performed by Statistical Package for Social Sciences Version 16.0 (SPSS, Chicago, IL, USA). Frequency differences between patients and controls were analyzed using chi-square test or Fisher’s test. P<0.05 were considered statistically significant.Results1. The total carriage frequencies of KIR2DL3, haplotype A and genotype A/A in Cushing’s disease patients with hyperprolactinemia were significantly higher than those in the control group (P=0.02, P=0.024, P=0.04).2. The carriage frequency of HLA-Cw*08 was significantly higher in patients compared with the controls (P=0.012).Conclusion1. The imbalance between inhibitory receptors and activating receptors may be influence the immune status. The immune status determined the prognosis and outcome of prolactinoma.2. The results demonstrated the importance of KIR and HLA-Cw genes on susceptibility to Cushing’s disease patients with hyperprolactinemia. The mechanisms will be of help in treatment strategies. Additional genetic and functional studies will be necessary to clarify the involvement of the mechanism in Cushing’s disease patients with hyperprolactinemia.