The Clinical Application Of Hydrogen Proton Magnetic Resonance Spectroscopy In Cerebral Gliomas

BackgroundGlioma is the most common primary intracranial tumor,it account for about 35.26%~ 60.96%of intracranial primary tumor.Because of its invasively growth,the outcome were poor even after all kinds of treatment for glioma patients,especially for high grade gliomas such as multiform gliolbastoma,rhey recurrent quickly with very poor prognosis.Nowadays glioma is a serious threat to human health and one of the most difficult malignant tumors for the neurosurgeon.People have reached a consensus on the treatment for gliomas:the principle of treatment for gliomas are resecting the tumor as much as possible and mostly protecting the nervous system function,then giving comprehensive treatment such as radiotherapy and chemotherapy individually after the operation.Surgical resection is always the primary treatment for gliomas;Meanwhile,it is necessary to consider the patients'quality of life while resecting the whole or sub-total of tumor.However,because of its infiltrative growth and unclear border, it is difficult to make total resction for gliomas.So get the precise border is very important for the glioma operation.Since there is no better means to determine the tumor boundaries,neurosurgeons still consider the enhanced tumor region on T1-weithted images as a reliable indicator for the border.The resection is also operated along the gliosis band between the enhanced tumor centre and the edeme tissue.However, the enhanced tumor border on MRI is not the real boundary of gliomas, it is only the representative of damage to the blood-brain barrier, There is still tumor cells infiltration in the peritumoral edeme zone.But the infiltrated neighboring region of brain tumor do not usually enhance on routine MRI,this will undoubtedly underestimate the extend of lesions.The high signal area on T2-weighted imaging are caused by the tumor cell infiltrationPartly,this method is easy to overestimate the tumor extent by the impact of the surrounding edema tissue.Conventional CT,MRI examination can only provide preliminary information for diagnosis about tumor location,size and shape.Doctors usually want to obtain much more comprehensive and accurate preoperative information about tumor grading,the extend of tumor infiltrating and so on.The in vivo magnetic resonance spectroscopy has been used more and more widely by clinical research in recent years,It can detect the metabolic changes of both tumor area and normal brain tissue non-invasively.It can make diagnosis and differential diagnosis for varying brain tumor by the metabolic concentration and ratios changes, analyse the malignance and grading for the tumor,evaluate the treatment outcome,detecte the recurrences and evaluate the prognosis for varying tumors.MRS imaging can find the difference between the normal tissue and primary or infiltrating tumor region in the fiber,analyse the changes in the tumor centre,the normal brain tissue and the edema zone.It plays an important role in observing the biological characteristics and determining the extent and nature of tumors.How to reach extensive resection and greatest protection of nerve function for glioma patients? How about the prospect of hydrogen proton magnetic resonance spectroscopy imaging in determining the border of glioma?There are to much research on differential diagnosis,malignance grading for glioma by using of magnetic resonance spectroscopy,but systemic research about using magnetic resonance spectroscopy to assess glioma infiltration and then to guide surgery and radiotherapy is rare both at home and abroad.The purpose of our experiment is prospectively analyzing the common spectroscopic metabolic ratios of glioma patients by using two-dimensional multi-voxel proton magnetic resonance spectroscopy scan sequence(2D-multi-voxel 1H-MRS),detecting the express of Ki-67 antigen in the different area of the post-operative pathological sample,and then investigating the application of magnetic resonance spectroscopy in evaluating glioma tumor cell invasion,tumor proliferation activity.Purposes of the researchA prospective study was performed on 16 cases of consecutive patients with suspected supratentorial hemispheric gliomas.All the patients underwent routine MR and MRS examination using uniform procedures,and then surgical resection within 72 hours of the MR examination.The common spectroscopy metabolic ratios were obtained from the enhanced tumor center,the peritumoral edema zone and the contralateral corresponding area by two-dimensional multi-voxel proton magnetic resonance spectroscopy imaging(2D-1HMRS) technology. Proliferative activity of the tumors in both the tumor center and the peritumoral edema zone were assessed by Ki-6? immunochemistry(Mb-1) on paraffin embedded tumor sections.Spectroscopic data was compared with Ki-67 labeling index,we also observe the histologic changes such as histological subtype,cellular atypia,cellular density in the different regions.By studing the correlation between spectroscopy metabolic ratios and the expression of ki-67 antigen in both the enhanced tumor center and peritumoral edema zone,we seek to explore the possibility of magnetic resonance spectroscopy in evaluating the biological behavior of invasion and proliferative activity in glioma patients.Materials and Methods1.objects of researchBetween January 2007 and April 2008,a prospective study was performed on 16 cases of consecutive patients with suspected supratentorial hemispheric gliomas.There were nine male and seven female patients, aging from 21 to 52 years old,the average age was 38.4 years old.All of the cases were confirmed by postoperative pathology.There were six low-grade(WHO gradeⅠ-Ⅱ) and ten high-grade tumors(WHO gradeⅢ-Ⅳ, multiform glioblastomas showed cystic or obvious necrosis were removed). All of the patients had no preoperative history of radiotherapy and chemotherapy,no previous intracranial inflammation or infection,no brain trauma and other related medical administration history.2.Magnetic resonance imaging examinationAll the patients had informed consent with the families,the spectroscopy data collection were carried out within 72h before the surgical resection.Here we use the signa 3.0T superconducting magnetic resonance scanning equipment by GE company.The routine MR T1 and T2-weighted image were completedby Spin-echo(SE) and fast spin-echo(FSE) sequences.The scanning parameters were as follows:T1WI(TR/TE,600/16ms),T2WI(TR/TE, 5100/138ms),thickness 5-8mm,Pitch 1.5mm.The point resolved spectroscopy(pointed-resolved spectroscopy sequence,PRESS) sequence was used for the multi-voxel two-dimensional magnetic resonance spectroscopy scanning.TR/TE:1000ms/144ms,voxel thickness 10mm, Pitch 2mm,NEX 1.The maximum lesion diameter on the T2WI axial imaging was selected as the reference.The field of spectral scanning(field of view,FOV) vary depending on the volume of the lesions,it should include the enhanced tumor center,the peritumoral edema zone and the contralateral corresponding normal brain tissue far away from the lesion itself.Phase matrix 160×160,the ROI(region of interest)were 1cm×1cm×1cm～1.5cm×1.5cm×1cm.The voxel positioning should avoid including the bone,blood vessels,cerebrospinal fluid,cerebral cortex,cystic necrosis,calcification and hematoma by tumor themselves.The shimming and water suppression were fulfilled automatically by pre-scanning.The enhanced regions on T1WI were defined as the tumor center,the peritumoral edema zone were defined as high signal on T2WI without enhancement.The spectroscopy data were acquired from the tumor center,the peritumoral edema zone about two centimeters away from the tumor center and the contralateral corresponding white matter respectively.In order to avoid the impact of paramagnetic contrast agent on the spectral data results,the spectroscopy acquisition were achieved before the enhancing scan as much as possible.3.Image post-processing and spectroscopy data acquisition The Sun sparc ADW4.0 image processing workstation and its own FuncTool image post-processing software were used to analyze the data.It can automatically obtain spectral peak map and metabolic-anatomic imaging at the same time.Regions of interest(ROI) were located in significantly enhanced tumor region(ROI-1),peritumoral edema zone 2cm away(ROI-2) and the normal brain tissue(ROI-3).The water signal was the internal reference,its chemical shift is 4.7ppm,NAA is located at 2.02ppm,Cr 3.02ppm and Cho 3.22ppm.Observe the metabolite concentrations distribution on the metabolic-anatomic map and the peak value on the spectroscopy map.The area under the curve representative the peak concentrations.Record the Cho/Cr,NAA/Cr and Cho/NAA ratios of the three ROI,the result were expressed in way of mean±standard deviation(x±s).4.Histology examinationDuring the surgical resection,we orient the direction by distinguishing the brain sulcus that can be seen clearly such as the superior frontal sulcus,the superior and inferior temporal sulcus,the sylvian fissure and so on.In order to ensure the spectral ROI and the pathological specimens were point-to-point consistent,we consecutively and completely resect the tumor and the peritumoral edema zone tissue.The surgical specimens obtained were immediately fixed with formalin,the thickness of paraffin-embedded sections were 4μm, conventional HE staining and Ki-67 antigen staining were finished.The results were judged single-blindly by skilled neurological pathologists.The cell with significantly nuclear atypia and Ki-67 staining positive were regarded as proliferative activity tumor cell.Six field by size of 33μm×33μm were randomly selected under the 200 times microscope,we count the highest density for tumor cells activity.Quote Stadlbauer'method,The ratio of positive tumor cells /total cells as a percentage was defined as the proliferative activity tumor cell density.5.Statistical analysisCalculate separately the average value of metabolic ratios in the three different regions,the results measured were expressed as(?)±s.SPSS13.0 statistical analysis software was used.Using repeated measure ANOVA,the means were further compared by LSD method;The differences between the tumor center and the edema zone were compared by paired-samples T test.The linear correlation were used to analyze the relationship between the spectroscopy metabolite ratio and the Ki-67 protein expression,we describe the correlation by using Pearson coefficients,the results were explained from ZOU.P＜0.05 was considered there were significant differences.Results1.The spectroscopy concentrations of different tumor regions.The spectroscopy peak map of the contralateral corresponding white matter were the same as normal brain tissue:NAA was the highest,Cr and Cho peaks were lower in different levels.The Cho concentration were increased significantly in both tumor center and the peritumoral edema zone,Cr and NAA were decreased,especially remarkable for the concentration of NAA.But from the tumor center,the peritumoral edema zone to the contralateral corresponding white matter,these changes do not always have a"gradual"nature.2.The spectroscopy metabolite ratios in different tumor regions.The spectroscopy metabolite ratios of NAA/Cr,Cho/Cr and Cho/NAA in the contralateral white matter region were 1.559±0.200,0.878±0.138,0.605±0.123 respectively;In the enhanced tumor center,the Cho/Cr and Cho/NAA ratios were increased to 2.212±0.566,2.880±0.677, the NAA/Cr ratios were decreased to 0.762±0.177;In the peritumoral edema zone,the ratios of Cho/Cr,Cho/NAA and NAA/Cr were 1.488±0.156,1.490±0.355 and 1.152±0.139 respectively.All of the metabolic ratios changed in the same trend as the tumor center. But from the enhanced tumor center,the peritumoral edema zone to the contralateral corresponding white matter regions,the ratios were not always changed step by step in the same way as the metabolic concentration.There were significant statistical differences among these metabolite ratios when compared with each other.3.Pathomorphological observation of tumor center and the peritumoral edema zone tissue.The conventional pathology were significant different between the tumor center and the peritumoral edema zone tissue under light microscope.The background of edema tissue weresparse with significantly reduced tumor cells density.The nuclear morphology such as atypia and mitosis in the edema zone were not so significant as the tumor center.There was no obvious necrosis in this region,we could also see different degrees of vascular endothelial cell proliferation.In addition,there seemed to be an obvious transition band between the enhanced tumor center and the peritumoral edema zone.4.The expression of Ki-67 protein.The ki-67 staining results in the postoperative pathologic Samples were shown as follows:the proliferative activity tumor cells density of the tumor center were7.87%±2.02%,while the peritumoral edema zone were 1.58%±0.44%;we could not see positive staining in the surgical resected edema tissue from traumatic brain injury patients.The difference Of ki-67 staining results betweent the enhanced tumor area and peritumoral edema zone were statistically significant.5.The correlation between the spectroscopy metabolite ratios and the expression of ki-67 protein.There was a positive correlation between the Cho/Cr ratios and the expression of ki-67 protein in the tumor center,the value of correlation coefficient was 0.603,but in the peritumoral edema zone this correlation did not exist,We could not see any relevance between the NAA/Cr ratios and the ki-67 protein expression both in the tumor center or in the peritumoral edema zone.The Cho/NAA ratios and the ki-67 protein levels were positive correlated in both the tumor center and the adjacent edema tissue.The Correlation coefficient were 0.842 and 0.514 respectively.Conclusions1.The pathological spectrum existed in the peritumoral edema zone suggested that there were infiltrating tumor cells in this area and the routine MRI can not define the accurate boundary of gliomas.The 2D-1HMRS can find new lesions that were not showed on routine MRI and have the potential of determining the accurate tumor cells infiltrating boundary.2.From the enhanced tumor center,the peritumoral edema zone to the contralateral corresponding white matter,the spectroscopic metabolic concentration and ratios do not always change step by step.The highest spectral concentration and ratios were not always positioned in the tumor center.The metabolism were more active in the edema zone in some patients.It suggest that glioma growth inhomogeneous,the MRS could provide a reliable targeted biopsy point for the pathological diagnosis.3.The tumor cells density,nuclear morphology index such as the nuclear size,atypia and mitosis in the peritumoral edema zone under the microscope were obviously different with the Enhanced tumor center.There seemed to be a transitional tissue band between these two regions.The difference of the Ki-67 protein staining between these two region were signicant,suggesting the different tumor cells proliferative activity.4.The Cho/Cr ratios and Ki-67 protein staining intensity were positive correlated in the enhanced tumor center,there were also positive correlation between the Cho/NAA ratio and Ki-67 protein staining both in the tumor center and the peritumoral edema zone,suggesting that the Cho/NAA ratios were more accurate for evaluating the biological properties of invasion and proliferation in gliomas.5.The infiltrating tumor boundary defined by spectroscopy metabolite ratios were not accurate,our research can not obtain the precise cut-off value to distinguish the tumor-infiltrating tissue from the normal brain tissue.These questions mentioned above still need further study.