| Pet dogs as the most common companion animals in modern society,their health and life has attracted a great attention.Tumor diseases have become a major life-threatening killer in dogs due to more carcinogenic stimulus.Tumors were pathological neoplasms caused bycell proliferation and differentiation process out of controlwith genetic mutation in animal bodies,can be divided into two basic types ofbenign and malignant.There are many factors affected the incidence of canine tumors,for instance,immune functiondecreased and endocrine disorders caused by breeds,age factorsand a variety of physical and chemical stimulation in feeding environment,what all can induced tumors.Canine mammary tumors(CMTs)is a primary type in female canine tumors,the clinical incidence of CMTs accounted for 50%or more of the total tumors,and half of CMTs are malignant.Spontaneous mammary tumors in dogs similar to human cancer in biologic and clinical features,which makes this animal model can be seen a good one in comparative medicine research.p53 and PTEN as the two tumor suppressor genes with highest mutation rate in human tumors have been study a lot.They have been used for detecting and diagnosing risk about numbers of human cancer,predicting prognosis and monitoring recurrence,but there is no report on the value of them as prognostic factors in CMTs.This study aims to analyze the incidence of clinical pet dogs' tumors,at the same time,focus on study the p53 and PTEN expression characters in CMTs,which is the highest incident tumor type.It's meaningful both in veterinary and human medicine.1.Clinical histopathological characterization and etiological analysis of tumors in pet dogsA total of 66 cases of clinical canine tumors were collected from some animal hospitals in Nanjing area.Result of blood routine examination in tumor dogs revealed lymphocyte increased but monocyte and neutrophils decreased frequently.The clinical appearances of benign and malignant patients are different:benign tumor generally has limited impact with body function,dogs won't show obvious systemic symptoms exceptthe focal mass.Butdue to metastasis,inflammation and other phenomena occur frequently in canine malignant tumors,affected dogs suffered from poor mental illness,diet and organ dysfunction.X-ray is an important diagnosisin the confirm the tumors of head,abdominal and surface.The results indicated that tumors were located in 8 tissues,mammary gland tumors and skin tumors were the two most common sites(45.5%,16.7%),followed by soft tissue(13.6%),oral(7.6%),penis and testis(6.1%),uterus and ovary(4.5%),lymphatic system(4.5%)and bladder(1.5%)tumors.The major population was small dogs(62.1%),Poodles had the highest tumors incidence(21.2%).Location and types of tumors varies between different breeds,CMTs can be seen often in small size dogs while soft and oral tumors happened frequently in middle or large size dogs.The dogs' age range from 10 months to 17 years,with the average age was 9.4 years.Results showed the middle and old groups were the major patients,but the number of young dogs increased.The onset age between breeds present a wave motion.Some special tumors can be influenced by dogs'gender and sterilization conditions.Diet conditions also related to tumors' incidence.2.Clinical histopathological analysis of mammary tumors in dogStatistical analyzed the relationship between the incidence of 30 CMTs cases and the influencing factors,then classified tumors by histopathology diagnosis.Results as followe:Small pure breed dogs as the dominant group in CMTs(90.0%),Poodle was the highest risk breed(33.3%).The age of dogs was mainly distributed in the 8-13 years old range(80.0%),cases below 4 years old or over 15 years old hadn't found.Tumor's type also can be affected by tumor size,the group which diameter belong 3 to 5 cm has the highest malignant rate(81.8%).There were 12 benign cases(40.0%)and 18 malignant cases(60.0%)found.Accord to the incidence from low to high in benign tumors were adenoma,fibroadenoma and intraductal papillary adenoma,what in malignant were intraductal papillary carcinoma,mixed caner and invasive carcinoma.Invasive carcinoma was the highest subtype(33.3%).3.Expression of p53 and PTEN mRNA in different type of canine mammary tumors25 samples(include 2 normal control mammary tissue)were divided into 4 groups(normal,benign,low stage-TNM ?-?,high stage-TNM IV/V)to detect the mRNA expression of p53 and PTEN in different CMTs by RT-PCR.The trend of p53 and PTEN mRNA expression with tumor malignance were seen as totally opposite.The expression level of p53 show up-regulared when samples with higher malignance,found the relative expression(RE)of p53 in all tumors groups were significant different from the normal tissues(P<0.05).Expression of p53 mRNA increased with the degree of malignancy mounted between groups.The median of RE in nonnal,benign,low stage,high stage groups were 0.96,2.45,2.58 and 3.79,respectively.A larger inter quartile range(IQR,2.06)in low stage group showed the expression were discreted.Significant differences can be seen between benign and high stage tissues or low stage and high stage tissues(P<0.05).On the other hand,transcript level of PTEN were decreased obviously in CMTs tissues relative to normal mammary tissues(P<0.05).The median RQ was 0.45 and the third quartile(Q3)was 0.67 in benign CMTs,which show the worst concentration condition in 4 groups.The median RQ declined in low stage group to 0.18,at the mean time,high stage group show the lowest expression level but its median RQ was higher than low stage group was 0.21,no difference between all tumor groups.4.Localization and expression of p53 and PTEN protein in different type of canine mammary tumorsp53 and PTEN protein expression were determined in 28 samples(2 normal control)grouped as aboved by Western Blot test.The results of immunoblotting show p53 has one specific band with 53 kDa and PTEN has one specific band with 54 kDa.Expression of p53 in CMTs was gradually increased compare to it's in normal tissues,and significant differences detected between all tumor groups and normal group(P<0.05).The relative quantity(RQ)of p53/?-actin in 3 tumor groupswas 2.73±0.33,2.93±0.37 and 4.01±0.39,respectively.The situation of PTEN was on the opposite with p53,the protein expression precipitly fall can be seen from normal to high groups(2.37 ±0.14,0.71 ±0.05,0.53 ±0.03 and 0.27±0.04,P<0.05).IHC was used to detected the localization of those two protein in 32 samples(2normal control and 30 CMTs tissues).p53 was found particularly localized in the nucleus,and the expression level of p53 was 62.5%,showed overexpressed in malignant tissues.PTEN was diffusely expressed in the cytoplasm,its expression lost can be seen with the malignancy increasing.Overall,56.2%of mammary tissues showed cytoplasmic PTEN loss.5.Relationship between p53 and PTEN expression and the clinical morbidity of canine mammary tumorsAnalyzed clinicopathological characteristics'correlation with p53 and PTEN expression.Results indicated tumor size has a significant relationship with the expression of p53,dogs whose tumor diameter among 3-5 cm has a highest p53 positive rate.Meanwhile PTEN showed a significant expression lost in cases of higher clinical stages.A strong negative correlation between the expression of p53 and PTEN was revealed by Spearman's rank correlation analysis(r=-0.367,P<0.05).Prognostic studies found that dogs with p53 negative expression or PTEN positive expression have a longer postoperative survival,respectively. |
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