Internal radiotherapy using 32P colloid or microsphere for refractory solid tumors.

OBJECTIVE: The aim of this work was to study the effectiveness of 32P colloids or microspheres, by arterial interventional administration or stromal injection in the treatment of refractory solid tumors. METHODS: By arterial intervention, under the guidance of computerized tomography, X-ray, ultrasonogram, or under direct vision of the surgical field, 32P microspheres (259-685 MBq) or radioactive colloid (281-666 MBq) was administered to 60 cases with refractory solid tumors. Tumor inhibition rate, side effects, survival period, and so on were observed. RESULTS: The tumor growth was obviously inhibited after the intratumoral injection of 32P colloid. The average survival time in the 60 cases was 35 months with a high tumor inhibition rate (93.4%). Thirty-one cases were completely relieved (51.7%), and 25 cases achieved partial remission (PR, 41.7%). One case with right lobe hepatocellular carcinoma has survived 90 months. The drug was ineffective only in four cases, including one patient who died of gastrointestinal hemorrhage and three of hepatic failure. No other obvious side effects were observed. Intratumoral necrosis, intense fibrosis in the tumor mass, and an integrated capsule encompassing the tumor were revealed by histological examination. CONCLUSIONS: Arterial interventional administration or stromal injection with 32P microspheres or colloid revealed a very fair clinical effectiveness in the treatment of refractory solid tumors. The range of safe effective dosage for 32P glass microspheres and 32P chromic phosphate in one treatment course is 555-740 MBq and 185-370 MBq, respectively.

[Expression of substance P in intestinal tissue and the relation between substance P and mucosa permeability in acute necrotizing pancreatitis in rat].

OBJECTIVE: To investigate the expression of substance P in the intestinal tissue and the relation between expression of substance P and pathological change of intestinal mucosa or mucosal Permeability in acute necrotizing pancreatitis (ANP) in rat. METHODS: One hundred and fifty adult Sprague-Dawley rats were randomly divided into sham operation group (n=50) and ANP group (n=100). In sham operation group, the pancreatic was only flipped over. In ANP group, ANP was produced by administration of 5% sodium taurocholate into the pancreatic duct. Rats were sacrificed at 0, 6, 12, 24 and 48 hours after reproduction of pancreatitis in both groups. The intestinal mucosal permeability was assessed with (99m)Technetium Diethylene Triamine Pentaacetic Acid method, and the expression of substance P was evaluated with reverse transcriptase polymerase chain reaction (RT-PCR) and Western blotting analysis. RESULTS: In ANP group, level of serum amylase raised at different time points after reproduction, intestinal mucosal permeability and the expression of substance P were also significantly higher than those in the sham operation group (P<0.01, respectively). The expression of substance P was positively correlated with mucosal pathological score (r=0.67, P<0.01) and intestinal mucosal permeability index (r=0.78, P<0.01) in ANP. CONCLUSION: The expression of the substance P and the permeability in intestine of rats are enhanced significantly in ANP. The expression of substance P is positively correlated with intestinal mucosal permeability.

[Construction and primary application of oligonucleotide microarray specialized for pancreatic adenocarcinoma-associated genes].

OBJECTIVE: To investigate the construction of oligonucleotide microarray specialized for pancreatic adenocarcinoma-associated genes and its application. METHODS: Pancreatic cancer related genes were purposely selected, and oligonucleotide microarray was prepared by spotting oligonucleotide probes onto glass slides coated with APS-PDC. Total RNA were extracted from frozen tissues with TRIzol method according to the manufacturer's protocol, and purified with QIAGEN RNeasy Kit. Labeled cDNA targets for hybridizations were synthesized by reverse transcription from control- and cancer-total RNA samples in the presence of Cy5-dCTP and Cy3-dCTP, respectively. The labeled probes were hybridized with oligonucleotide microarray for 16 h to 18 h. Hybridized microarray was scanned by Agilent laser scanner, and the acquired image was analyzed by Imagene3.0 software. The intensity ratio of Cy3 and Cy5 were calculated. To confirm the expression profiles of these genes, quantitative reverse transcription-PCR (Q RT-PCR) was carried out with CDC25B and TUSC3 genes. The product of PCR were quantitated by comparative Ct method. RESULTS: The signal of microarray hybridization was clear, and the images had a lower background and higher signal-noise ratio. The signal of positive control spots were uniform, and spots of negative control and blank signal were fairly low. In comparison with normal pancreas, 24 differential expressed genes were identified, which included 17 up-regulated and 7 down-regulated genes. The results of Q RT-PCR demonstrated that the expression of CDC25B and TUSC3 in pancreatic cancer were increased and decreased respectively, which consistent with microarray hybridization. CONCLUSIONS: The oligonucleotide microarray specialized for pancreatic cancer are desirable for its specialty, flexibility and sensitivity, which can simultaneously and parallelly detect multiple pancreatic cancer-associated genes. In contrast to normal pancreatic tissues, the genes expression profile are different significantly in pancreatic cancer.

[Role of neurokinin-1 receptor in lung injury in rats with acute necrotizing pancreatitis].

OBJECTIVE: To investigate the expression of neurokinin-1 receptor (NK-1R) in the lung tissue, and the relationship between expression of NK-1R and lung injury in rats with acute necrotizing pancreatitis (ANP). METHODS: One hundred and twenty adult Sprague-Dawley rats were randomly divided into ANP and control groups. Animals in group ANP were induced by the retrograde intraductal infusion of 5% sodium taurocholate (0.1 ml/kg), and animals in normal control group received laparotomy only. The accumulation of polymorphonuclear leukocytes in lung tissues was measured with myeloperoxidase (MPO) assay. Lung endothelial barrier destruction was measured by lung capillary permeability (LCP). Reverse transcription polymerase chain reaction (RT-PCR) was used to determine the mRNA expression of NK-1R, western blot analysis was used to determine NK-1R protein expression levels, and immunohistochemistry was used to localize expression site of NK-1R. RESULTS: NK-1R mRNA level was enhanced in the lung of ANP compared with normal control group. Western blot analysis showed overexpression of NK-1R protein level exited in ANP group. Statistical analysis revealed correlation between NK-1R mRNA and MPO (r=0.83, P<0.01) and LCP (r=0.79, P<0.01) respectively. With immunohistochemistry staining, moderate to strong NK-1R immunoreactivity was localized to alveolar membrane, I epithelium, II epithelium and polymorphonuclear leukocytes in the lung of ANP. CONCLUSION: In ANP, overexpression of NK-1R contributes to disturbance of neuropeptides loop, resulting in aggregation of neutrophilic granulocyte and promoting deterioration of lung injury.

[Gene expression analysis in liver tissue at a single cell level by nested polymerase chain reaction and laser microdissection].

OBJECTIVE: To investigate the measurements of gene expressing at a single hepatocyte level. METHODS: Individual hepatocyte was isolated from cryostat tissue section using laser microdissection technique. To detect the mRNA expressed by single hepatocyte, RNA was extracted, reversely transcribed to cDNA and amplified by nested polymerase chain reaction (PCR). RESULTS: Single cell was microdissected from cryostat tissue using an ultraviolet laser micromanipulator. The RNA could be extracted from the isolated cell(s), and the RT-PCR production could be observed after electrophoresis, whose quantitation was compatible with the number of cells. CONCLUSION: Combining laser microdissection and nested RT-PCR can monitor gene expression at a single cell level in vivo.

Relationship between overexpression of NK-1R, NK-2R and intestinal mucosal damage in acute necrotizing pancreatitis.

AIM: To study the expression of neurokinin-1 receptor (NK-1R) and neurokinin-2 receptor (NK-2R) in distal ileum of acute necrotizing pancreatitis (ANP) and to evaluate the relationship between expression of these two receptors and intestinal mucosal damage. METHODS: A total of 130 adult Sprague-Dawley rats were randomly divided into two groups: the rats in ANP group (n=80) were induced by the retrograde intraductal infusion of 30 g.L(-1) sodium taurocholate. And the rats in normal control group (n=50) received laparotomy only. Sacrifices were made 6 h, 12 h, 24 h and 48 h later in ANP and normal control group after induction respectively. Intestinal mucosal permeability was studied by intrajejunal injection of 1.5 mCi radioactive isotope (99m)Tc-diethlene triamine pentacetic acid (DTPA) and the radioactivity of (99m)Tc-DTPA content in urine was measured 6 h, 12 h, 24 h and 48 h after induction. Then the pancreas and intestine were prepared for pathology. Reverse transcription polymerase chain reaction (RT-PCR) was used to determine the mRNA expression of NK-1R and NK-2R, and Western blot was used to investigate the protein level of NK-1R and NK-2R. RESULTS: In ANP rats, serious histologic damages in intestinal mucosa were observed, and the radioactivity of (99m)Tc-DTPA in urine increased significantly in the ANP group. RT-PCR revealed that NK-1R and NK-2R mRNA level was overexpressed in the distal ileum of ANP as compared with the normal control group. Western blot discovered stronger NK-1R (14-fold increase) and NK-2R (9-fold increase) immunoreactivity in the intestinal mucosa of ANP rats. Moreover, the overexpression of NK-1R was associated with mucosal pathological score (r=0.77, P<0.01) and intestinal permeability (r=0.68, P<0.01) in ANP rats. CONCLUSION: NK-1R and NK-2R contribute to disrupted neuropeptides loop balance, deteriorate intestinal damage, and are involved in pathophysiological changes in ANP.

[Laser microdissection of a single cell from colon tissue for gene analysis].

AIM: To investigate the method of detecting gene expression in colon tissue at a single cell level. METHODS: Individual cell(s) were picked up from colon frozen section using laser microdissection. RNA was extracted, reverse transcribed to complementary DNA (cDNA). cDNA was then analyzed by nested reverse transcription polymerase chain reaction (nested RT-PCR) using two pairs of primers. RESULTS: Single cell(s) were selectively picked up using an ultraviolet laser micromanipulator. RNA was extracted, reverse transcribed and used for nested RT-PCR. Amplification products of cDNA from down to a single cell could be clearly visualized in the agarose gel. CONCLUSION: The combined utilization of laser microdissection and nested RT-PCR provides an opportunity to analyze gene expression at single cell(s) level in colon tissue.