Experimental Studies On Tumoral Angiogenesis Of Rat C6 Gliomas With Perfusion Weighted Magnetic Resonance Imaging

Objective Perfusion-weighted MRI (PWI) was performed in implanted C6 rat gliomas to analyze its features in PWI and evaluate tumor vascularity by correlating with immunohistochemical findings(microvascular density, MVD). Materials and methods 10 male SD rats were implanted of C6 glioma cells at right nucleus caudatus through a stereotactic apparatus. 4 weeks later, the rats were imaged with conventional MRI and PWI in a 3.0-T GE MR scanner. Maximal diameters of tumors were measured in enhanced T|W images. The original images of PWI acquired from scanner were transmitted to the workstation. Perfusion software was used to create perfusion maps(rCBV, rCBF and MTT) and graphs for analysis. The rats were killed and their brains were removed within 24 h after MRI. Each rat brain was examined histologically using HE and immunohistochemical staining for FVIII. The histological features of gliomas were observed and MVD of each tumor was counted manually using a light microscope. The rrCBV, rr CBF and MTT values ,as well as the counted MVD numbers were input into a personal computer for statistical measurement. Pearson regression or correlation software was used to determine the relations between rrCBV, rrCBF and MVD respectively. Correlation coefficient was also calculated and t-test was made (with statistical significance when P < 0.05). Results Gliomas with different size were viewed growing in each rat brain 4 weeks later after tumor implantation. The CBV, CBF of gliomas were significantly higher than those found in healthy contralateral brain tissue . The signal intensity of tumors declined deeper than that of contralateral brain tissue during the first pass of contrast medium and the recovery time prolongs. Some of lesions created irregular first-pass decline curves. Although the MTT of tumors slightly prolonged, no significant difference existed between tumors and contralateral brain tissue(P> 0.05).Immunohistochemical staining of 10 tumors for FVIII all positively expressed in endothelial cells of vessels which present brown irregular spots or stripes. Correlation between rrCBV , rrCBF of tumor core and microvascular density were present (r = 0.861, P = 0.001, r = 0.731, P = 0.016 respectively). Conclusion Both rCBV and rr CBF of the solid tumor component are well correlated with MVD of C6 gliomas. MR PWI can evaluate tumoral microvessel in vivo.Part II A Preliminary Study on Functional Response of Tumor Vasculature to Hypercarbia by Perfusion wighted MR ImagingAbstractObjective The purpose of the study is to analyze how valuable of MR PWI in judging maturity and variation of tumor vasculature by means of studying changes of MR PWI values in normal rat brain tissue and C6 gliomas under conditions of hypercarbia ( increase of PaCO₂) and correlating with results of histological staining (smooth mussels actin, SMA). Materials and Methods 20 male SD rats were randomly assigned into 2 groups: tumor and control group. 10 rats of tumor group were implanted of C6 glioma cells at right nucleus caudatus through a stereotactic apparatus and those of control group were injected normal saline at the same region. 4 weeks after tumor implantation, the rats of both groups were imaged with MR PWI in a 3.0-T GE MR scanner pre- and post- (15 minutes delay) inhalation of a mixture of 10% CO₂ and 90% air. The original images of PWI acquired from scanner were transmitted to the workstation. Perfusion software was used to create perfusion maps(rCBV, rCBF and MTT) and graphs for analysis. PaCO₂ and blood PH values were monitored before and after mixed gas inhalation. The rats of tumor group were killed and their brains were removed once the MRI scan over. Each rat brain was examined histologically using HE and immunohistochemical staining for SMA. The histological features of gliomas were observed and SMA positively stained vessels of each tumor was counted manually using a light microscope. The PWI values , the counted number of SMA positively stained vessels, as well as the PaCO₂ and blood PH values were input into a personal computer for statistical treatment. The significance between changing rate(%) of perfusion values at right nucleus caudatusin normal rats(control group) and that at left ones in rats of tumor group was calculated using grouping two-tailed t-test. The difference between changing rate(%) of rCBV and rCBF mixed gas inhalation of normal rat brain tissue and that of tumor core was determined using paired single-tailed t-test. The difference between the number of SMA(+) within gliomas and that of controlateral brain tissue was calculated using F test. Pearson regression or correlation software was used to determine the relations between changing rate of rCBV & rCBF after mixed gas inhalation and number of SMA positively stained vessels respectively. Correlation coefficient was also calculated and t-test was made. Results Gliomas with different size were viewed growing in each rat brain 4 weeks later after tumor implantation. After mixed gas inhalation, PH decreased and PaCO₂ increased significantly(P< 0.025 and 0.001 respectively). No significant difference of blood gas value changes exists in tumor group and control group(P > 0.5). The rCBV and rCBF values of tumor core are remarkably greater than those of contralateral brain tissue. The increasing rate of rCBV and rCBF values in gliomas is significantly less than those in normal brain tissue, especially of rCBV(t = 5.05, P < 0.001 and t = 2.355, P = 0.021). Immunohistochemical staining of 10 tumors as well as contralateral brain tissue for SMA all positively expressed in smooth mussels of vessels which present brown regular loops or tubes.The vascular walls of SMA(+) labeled vessels within tumors are thinner and the vascular lumen of tumors wider and smaller in number (t = 1.86, P < 0.001) than those within normal brain tissues. Correlation between changing rate of rCBV , rCBF of tumor core and number of SMA positively stained vessels were present (r = 0.504, P = 0.137, r = 0.607, P = 0.062 respectively), while changing rate of rCBV , rCBF in normal brain tissue correlated well with number of SMA positively stained vessels(r = 0.721 and 0.525, . P < 0.05).Conclusion Hemodynamic changes in both gliomas and normal brain tissue following PaCO₂ changes can be reflected by MR PWI. Although it is difficult for PWI to completely qualify mature vessels within tumor core currently, maturity of tumor vessels could be indirectly revealed (especially some variation of tumor vasculature) by this technique.Objective To observe the theropeutic particularly the antiangiogenic effect of Stereotatic Radiosurgery (SRS) on rats C6 glioma and to study the practical value of PWI in evaluating the response of the tumor to this treatment. Materials and Methods 20 male SD rats were implanted of C6 glioma cells at right nucleus caudatus through a stereotactic apparatus. The rats were randomly assigned into 2 groups with 10 in group A(without any theropy) and group B (with SRS treated) each. The rats in group B were performed SRS therapy at the center of tumors 4 weeks later after glioma cells implantation. PWI was performed to measure the regional cerebral blood volume (rCBV) and regional cerebral blood flow (rCBF) of tumors before therapy at both groups and 1 week later after therapy at group B. MR PWI was performed in a 3.0-T GE MR scanner pre- and post- (15 minutes delay) inhalation of a mixture of 10% CO₂ and 90% air. The original images of PWI acquired from scanner were transmitted to the workstation. Perfusion software was used to create perfusion maps(rCBV, rCBF and MTT) and graphs for analysis. PaCO₂ and blood PH values were monitored before and after mixed gas inhalation. The rats were killed and their brains were removed at the entire end of MRI. Each rat brain was examined histologically using HE and immunohistochemical staining for FVIII and SMA. The measured PWI values , the counted number of SMA positively stained vessels, as well as the PaCO₂ and blood PH values were all input into a personal computer for statistical analysis. The significance between rrCBV & rrCBF values , increase rate(%) of rCBV & rCBF in hypercarbia before treatment and those after therapy using paired single-tailed t-test respectively. The difference between MVD & the number of SMA positively stained vessels[SMA(+)] of group A and those of SRS treated group B. SMA(+)/MVD (%)of each rat tumor was calculated. The difference between SMA(+)/MVD (%) value of group A and that of treated group B. Results Gliomas with different size were observed growing in each rat brain 4 weeks later after tumor implantation. There was no significant difference in average tumor maximal diameter between group A and B (t = 0.132, P>0.5 ) . The tumors of group B remarkably decreased in volume and obvious necrotic area appeared 1 week later after SRS therapy. Observed through light microscope, the density of glioma cellsdecreased because of multiple cystic necrotic areas. After mixed gas inhalation, PH decreased and PaCO₂ increased significantly. No significant difference of blood gas value changes existed in two groups (t = 0.224 and 0.4 respectively, P > 0.5). The rrCBV and rrCBF of tumor core in group B declined greatly after SRS treatment (rrCBV: t =7.478 , P = 0; rrCBF: t =10.13, P = 0) though still higher than controlateral brain tissue. The necrotic area showed "black "in rCBV and rCBF maps for no blood perfused inside. The changing rate of rCBV and rCBF after hypercarbia in group B increased greatly (t = 12.27 ,P = 0; t = 6.364 ,P = 0.00013 respectively). Immunohistochemical staining of 20 gliomas for FVIII and SMA all positively expressed. The MVD of tumors in treated group B is statistically less than that in group A(t = 3.466, P=0.001) while the number of SMA positively stained vessels in in treated group B is nearly equal to that in group A (t = 0.513, P = 0.614 ) . The SMA(+)/MVD (%) in tumor of group A is significantly lower than that of SRS treated group B (t = 2.373, P = 0.014 ) , which the percentage of vessels with smooth mussel cells in treated group B is much higher than that of group A. Conclusion SRS could inhibit the angiogenesis of rat C6 glioma while "mature" vessels with smooth mussel cells are insensitive to SRS. MR PWI could detect the efficiency of antiangiogenic therapy and reflect variation of tumor vasculature to some extent by means of changing PaCO₂...