P21^WAF1/Cip1, an inhibitor of cyclin-dependent kinases, is a critical downstream effector in the P53-specfic pathway of growth control. Increased expression of P21^WAF1/Cip1 has been found to reflect the status of the P53 tumor-suppressor pathway. We investigated the expression of P21^WAF1/Cip1 in a relatively small, but well-characterized group consisting of 28 hepatocellular carcinomas. The samples were previously studied for P53 gene mutation. P21^WAF1/Cip1 expression were identified byin situ hybridization and immunohistochemistry. Positive ISH for P21^WAF1/Cip1 transcripts was found in 18 of 28 cases (64. 3 %). All positive cases by ISH showed detectable P21^WAF1/Cip1 protein reactivity by IHC. No relationship was found between p21^WAF1/Cip1 staining and P53 mutational status. No associations were seen with tumor metastasis, size and tumor grade, except for tumor differentiation status which showed higher frequency of P21^WAF1/Cip1 expression in moderate-well differentiated HCCs than poorly differentiated tumors (P< 0. 05). It is concluded that expression of P21^WAF1/Cip1 is common in HCCs, but does not correlate with P53 mutational status or pathological parameters investigated except for tumor differentiation. Also, there may be other factors beside P53 that regulate P21^WAF1/Cip1 gene expression in HCCs.

The effects of chimeric anti-CD4 human/murine chimeric antibody and murine anti-CD4 monoclonal antibody (McAb) on the proliferation induced by anti-CD3 McAb, phytohemagglutinin (PHA), IL-2, and allogeneic cells were studied. The results showed that chimeric anti-CD4 antibody and murine anti-CD4 McAb could inhibit the proliferation induced by the above inducers and the inhibitory effects were related to the dosage of the antibodies.

Mouse B7-1 cDNA was cloned by RT-PCR from BALB/C mouse splenic cells and inserted into pcDNA3 to construct an eukaryotic expression vector. This constructor was named pCD-mB7-1. in which the B7-1 cDNA was identified to be consistent with the data from other researchers. pCD-mB7-1 plasmid was transfected into B16(F0) cells, and effective expression of mB7-1 in these tumor cells could be detected till the 6th month by RT-PCR and RNA hybridization. Specific cytotoxity assay of lymphocytes was conducted after culturing with tumor cells and the results demonstrated that B16 cells transfected with B7-1 gene were more effective than B16-wt and B16-neo in inducing specific cytotoxity of lymphocytes against B16-wt cells. It is suggested that expression of B7-1 gene in tumor cells could enhance the immunogenicity and induce the effective antitumor immunity.

In order to measure the plasma levels of interleukin-4 (IL-4). cyclic adenosine monophosphate (cAMP) and cyclic guanosine monophosphate (cGMP) in patients with chronic obstructive pulmonary disease (COPD) and observe the effects of oral theophylline on them, we divided 28 COPD patients into COPD experimental group and COPD control group. Plasma levels of IL-4, cAMP and cGMP as well as parameters of pulmonary function tests were measured in these 2 groups before and after 2 weeks of treatment with oral theophylline (Protheo, 400 mg, qd) or placebo. Plasma levels of IL-4 and cGMP were significantly elevated in patients with COPD as compared with normal controls (P< 0. 05), while cAMP and cAMP/cGMP were significantly lower than those in controls (P < 0. 01). Plasma level of IL-4 was inversely correlated with forced expiratory volume at the first second (FEV₁) and with maximum expiratory flow rate at 50 % of forced vital capacity (V₅₀) (bothr= −0.46,P< 0.05) while it was directly correlated with the scores of the clinical manifestations (r=0.57,P< 0.05) in COPD patients. Two weeks after treatment with theophylline, IL-4 and cGMP in COPD experimental group were decreased significantly while cAMP and cAMP/ cGMP increased significantly (P< 0. 05). The change of IL-4 was inversely correlated with the changes of FEV₁, and V₅₀(r=−0.53 and -0.54, respectively,P< 0. 05). Two weeks after placebo treatment, the COPD control group did not show such changes. We are led to conclude that IL-4 might play a role in the pathogenesis of the airway inflammation and air flow limitation in COPD patients and the mechanisms of theophylline’s therapeutic efrects of attenuating air-flow limitation may partially depend on its anti-inflammatory effects on the airways which, in turn, is dependent on its inhibitory effects on some inflammatory mediators such as IL-4.

In order to investigate the regulating role of Astragapolysaccharide (APS) in the mice model of asthmatic airway inflammation, the airway eosinophil number, spleen T lymphocyte proliferation, level of IL-2 production and their relationships were studied in sensitized mice and sensitized mice treated with different concentrations of APS. The results showed that the number of eosinophils as well as lymphocytes in the airway of the sensitized animals were significantly increased, and a marked positive correlation between the inflammation cells and spleen T lymphocyte proliferation was found. Moreover, there was a positive correlation between inflammation cells and the level of IL-2 production. The APS of given dosage could significantly reduce the number of eosinophils in the airway of the sensitized animals. At the same time the level of IL-2 secreted by spleen T lymphocytes stimulated with ConA was also significantly decreased and there was a marked positive correlation between them. Our results suggested that APS of given dosage could prevent antigen-induced the number of eosinophils infiltrating into the airway of sensitized mice and inhibit the proliferation and activation of lymphocyte and IL- 2 production. Through its immuno-regulating effect, APS can be helpful in the treatment of asthma.

In order to study the cellular origin of muscularization in non-muscular arterioles of the lung, the pulmonary vascular pericytes-culture was established. The terminal lung tissue of the rat was taken out and minced. Then 0.5 % of type IV collagenase solution was added for digestion and the microvascular segments were obtained by screening. The targeted cells were cultured by “selective conditioned media”. Under phase-contrast microscope, the cultured cells were large in size with ragged margin and numerous pseudopodia, which imparted tubule-like structure. There was no contact inhibition in growing cells, so multiple layers developed. When they were confluent, there were morphologically no “hillock and dale” growth pattern as in smooth muscle cells or “weave-like” pattern as in fibroblasts. The ultrastructure of cultured cells showed numerous digital processes, moderate amount of rough and smooth endoplasmic reticulum, rich Golgi’s apparatus, microfilaments, few lysosomes without myofilaments and dense bodies. Immunohistochemical staining revealed that the cultured pericytes had same kind of cellular skeletal protein, α-SM-actin, like smooth muscle cells. The cultured cells also exhibited positive reaction to CD₃₄ antigen and S-100 antigen, which were negative in smooth muscle cells and fibroblasts. The cell growth pattern, ultrastructure and immunological phenotype suggested that the cultured cells had characteristics of vascular pericytes. Pericytes are one of the components of microvascular cells, and the establishment ofin vitro culture technique of pericytes is of significance for further exploration of the muscularization of non-muscular arterioles in lung and the mechanism of structural remodeling of pulmonary vessels.

To understand P53 gene change of non-Hodgkin’s lymphoma (NHL) and human malignant lymphoma cell lines, the exons 5–7 of 29 patients with NHL and 9 kinds of human malignant lymphoma cell lines were studied by silver staining PCR-SSCP technique. Three cases of P53 gene point mutation was found in 29 cases of NHL. Mutation developed in exon 5 in 2 cases, and in exon 6 in 1 case. They were all diffuse lymphoma. In mutation cases, B-cell lymphoma accounted for 2 cases and the other one was T-cell lymphoma. There was no P53 gene mutation in low-grade follicular lymphoma. Seven strains out of 9 kinds of lymphoma cell lines had P53 gene point mutation. One strain had the mutation in exon 5; 5 strains in exon 6 and 1 strain in exons 5, 6, 7. There was a high mutation rate in lymphoma cell lines and low mutation rate in NHL patients. P53 gene plays an important role in lymphoma cell line establishment, cell regeneration and disease evolution.

Mice with immune-induced aplastic anemia (AA) were given 5 mg ligustrazine intraperitoneally twice a day. On the 14th day, the expression of CD_49d CD_49c cyclinD₂ in bone marrow mononuclear cells (MNC) was examined by flow cytometry, and VCAM-1 on stromal cells was immunohistochemically measured by Strept Avidin-Biotin Complex (SABC). The expression of CD_49d, CD_49c, VCAM-1 and cyclinD₂ in ligustrazine-treated group was significantly higher than that in AA group (P< 0. 01), but the ratio of G₀+G₁ phase cells was significantly lower than that in A A group (P< 0. 01). The results showed that ligustrazine could improve the expression of adherent molecule and cyclin D₂ in the bone marrow of mice with immune-induced aplastic anemia, thereby promoting the growth of hematopoietic cells.

To evaluate the effect ofSalvia Miltiorrhiza on coronary collateral circulation in dogs with experimental acute myocardial infarction, 12 dogs were randomly divided intoSalvia Miltiorrhiza group (group D) and control group (group C). Acute myocardial infarction model was established in all dogs,Salvia Miltiorrhiza or saline was administered to the 2 groups respectively for 21 days. Coronary angiography was performed before and after ligation of coronary artery. The capillaries and distribution vessel density in tissue sections of myocardia were assessed. Angiography 2 h and 22 days after ligation of coronary artery revealed significant augmentation of collateral vessels, and capillaries and distribution vessel density in ischemic zone on day 22 showed statistically significant augmentation in group D as compared with that of controls (P< 0. 05 andP< 0. 01). It is concluded thatSalvia Miltiorrhiza might improve the opening and formation of coronary collateral circulation of ischemic myocardia in dogs and thereby protecting myocardia from ischemia.

In order to assess the value of Doppler tissue imaging (DTI) in detecting viable hibernating myocardium, 20 patients with coronary artery disease and chronic left ventricular dysfunction underwent low dose dobutamine stress e-chocardiography and low dose dobutamine stress DTI. The results showed that among the 100 asynergic segments, 35 segments showed improvement after dobutamine infusion (group H) and no changes were observed in the remaining 65 segments (group N). The left ventricular echocardiographic score index decreased from 1.60±0.35 to 1.44±0.36 (n = 20,P< 0.01). During low dose dobutamine stress DTI. there was no difference in the values of velosity of S wave (V₃) before dobutamine infusion between two groups. However, after dobutamine infusion, the values of V₃ and VR in group H were significantly higher than those in group N (V₃:10.1±3.0 cm/s vs 7. 3±2. 2cm/s,P< 0.01; VR: 60 %±41% vs 25%± 32%,P< 0. 001). 95.7% asynergic myocardial segments with VR≤0 had no viability while 86% asynergic segments with VR> 80% were viable myocardium. It is concluded that the different reactions to dobutamine stress between hibernating and necrosis myocardium could be showed by DTI and it is more clinically significant when VR≤0 and VR> 80%.

One hundred and twelve cases of familial myasthenia gravis (MG) from 44 families selected from 2100 patients with MG diagnosed since 1983 in the Department of Neurology were studied. The clinical pictures and immunological features of the patients showed a great resemblance to those of sporadic cases. The pedigree analysis disclosed that the hereditary patterns of familial patients were basically Mendellian autosomal inheritance. Many predisposing factors such as fever, infection, use of aminoglycoside or vaccines, played an important role in presenting the phenotype of subclinical cases. The HLA genotyping suggested that the complement polymorphism C4A * 4, the complotype S42, and the genes 0901 and 1301 of DRB1 allele, were related to the pathogenesis of MG. It was concluded that the phenotype of MG may be the result of interaction between hereditary defects and environmental factors.

Wilson disease (WD) is an autosomal recessive disorder of copper metabolism. To establish an efficient, accurate and fast diagnostic method for carrier detection and presymptomatic identification of WD in Chinese population, we studied haplotypes of short tandem repeat (STR) polymorphisms flanking the WD gene in 40 Chinese WD families. The results suggested that this genetic diagnosis system based on the four STR polymorphisms is of high value for the detection of potential carriers and WD homozygotes in families with at least one previously affected child. It is an efficient, accurate and fast diagnostic method that can be well suited for routine use in clinical laboratories.

To investigate the significance of hepatocyte apoptosis, proliferation and oncogene c-fos expression in carbon tetrachloride (CCl₄)-induced cirrhotic rat liver. Rat cirrhosis was induced by subcutaneous injection of 50% (v:v 1:1) CCl₄. Hepatocyte apoptosis, proliferation and oncogene c-fos expression were examined with TUNEL, PCNA and c-fos immunohistochemical methods in control group and treatment group 72 h, 5, 7, 11 and 15 weeks after CCl₄, induction. Hepatocyte apoptosis was rarely seen in control rat liver. The hepatoeyte apoptosis was obviously increased 72 h after treatment. Fifteen weeks after treatment, the apoptosis was still more obvious in treatment group than that in controls. PCNA was constantly expressed in CCl₄ group, with highest level at middle phase. C-fos was positive 7 and 11 weeks after CCl₄ treatment. The results suggest that: 1) apoptosis is involved in rat liver damage at the early phase in CCl₄-induced injury, and the process can alleviate nodule reconstruction or eradicate potentially mutational hepatocytes at the later phase; 2) hepatocytes constantly proliferate in CCl₄-induced rat liver cirrhosis, especially at the middle phase; 3) c-fos might modulate hepatocyte proliferation in CCl₄-induced rat liver cirrhosis.

To evaluate the effect of lower intensity anticoagulation therapy in patients with mechanical prosthetic valves, laboratory-based hematological assays including prothrombin time (PT), activity of factor X, antithrombinI (ATI), D-dimer, fibrinogen (Fg) and platel et al pha-granular membrane protein (GMP-140) were performed in 65 patients who had been on warfarin treatment for over one month. The patients were assigned to 3 groups on the basis of their International Normalized Ratios (INR), ranging from 2.00 to 2.50; 2.51 to 3.00; 3.01 to 4.50, respectively. The results showed that the D-dimer, Fg, GMP-140 levels were higher after mechanical valve replacement than those before operation, indicating, the activation of coagulation and fibrinolysis system and the damage of platelets. Lower intensity anticoagulation therapy (INR 2.00 to 2.50) could effectively inhibit the activity of factor X and increase the level of ATI. There were no appreciable differences among D-dimer, Fg, GMP-140 and ATI in the 3 anticoagulation intensity groups. These results suggest that in patients with new generation mechanical prosthetic valves, target anticoagulation level (INR 2.00 to 2.50) may result in good protection from thrombo-embolism.

Serum anticardiolipin antibody (ACA) was measured in 91 patients with cerebral infarction (CI), 42 patients with cerebral hemorrhage (CH) and 30 healthy controls. The results showed that the ACA in CI and CH patients was significantly higher than in controls and IgG-ACA was the most important iso-type. Stroke in ACA positive group tended to be recurrent and of multi-focuses. Positive rate of IgG-ACA reached its peak within the first week after stroke on-set. The results suggested that ACA was an independent risk factor in CI and CH and might be valuable in stroke prediction.

Ten girls with Turner syndrome were treated with a combination therapy of recombinant human growth hormone (R-hGH) and low dose stanozolol for a period of 8 to 36 months. The results showed that when compared with the growth rate before the treatment, the growth rates after treatment with R-hGH and stanozolol showed a sustained increase, reaching 9. 0±l.9 cm/year during the first year of treatment; the height age increase by 2. 5±0.8 years while the bone age increase were 1. 0±0. 7 years; and the predicted final adult height at the end of the first year of the treatment increased to 149. 4±6. 1 cm compared to their original mean of 142. 8 ±4. 2 cm. We are led to conclude that therapy with R-hGH in combination with stanozolol can increase the growth velocity and significantly increase the predicted adult height of children with Turner syndrome.

To compare Th2 cytokines levels in livers and bone marrow of mice infected withSchistosomiasis Japonica and understand their implication, ABC immunohistochemical staining technique and multimedia pathologic pictures analysis were used to detect the IL-4, IL-5 and IL-10 derived from livers and bone marrow of infected mice. Results showed that in the livers of infected mice, IL-4, IL-5 and IL-10 were obviously increased over time. The level of IL-4 was increased most among the 3 indicators. In bone marrow of infected mice, however, even though IL-4 was slowly increased over time, it was lower than that of livers in 10th and 12th week. The level of IL-5 was higher than that of livers during 12th weeks. Before 10th weeks IL-10 was elevated. After 12th week IL-10 was decreased and obviously lower than that of livers. It is concluded that in mice infected withSchistosomiasis Japonica, the immune response occurred mainly in the liver. Therefore, in bone marrow, IL-5 showed obvious auto-secretion. It might be related to its biological ability to induce B cell division and eosinophil differentiation.

The direct immunogold silver staining (D-IGSS) method was used to detect the viral antigen in the extremity blood of 67 cases of hemorrhagic fever with renal syndrome. The positive rate of viral antigen was the highest during the fever, hypotension and oligouria phrase; and the rate dropped gradually during the polyuria and convalescent phase. It is suggested that clinical staging was positively related with the percentage of the viral antigen positive cells (P< 0. 001). It is concluded that the positive rate was related to the extent of the injuries by direct viral attack and immune reaction. The D-IGSS was proved to be fast, simple, economical, with high sensitivity and specificity.

To explore the changes in MR findings of the bone marrow of vertebral bodies after radiation therapy, 55 subjects who had been on irradiation treatment were studied. Irradiation dosage ranged from 2100 cGy to 7500 cGy. The interval between radiotherapy and MR examination varied from 7 weeks to 20 years. On T1WI images (T1WI) signal intensity increase of the bone marrow of those vertebral bodies was apparent. The areas with altered signal intensity rendered well-defined margins corresponding to the field of irradiation. The similar finding was found on T2WI. On STIR or T1WI with fat suppression, the signal intensity of the areas was decreased. It is concluded that the changes of signal intensity on SE or STIR sequence is believed to be secondary to diffuse fatty infiltration and the results of replacement of the normal bone marrow by relatively increased yellow bone marrow.

In order to understand the roles of nitric oxide (NO) in pulpalgia and pulpitis, the histochemistry of nitric oxide synthase (NOS) in the neurons of trigeminal ganglion in experimental pulpitis rat and human inflammatory dental pulp tissues were histochemically studied by NADPH-diaphorase (NADPH-D) techniques. Results showed that NADPH-D positive neurons were scattered in rat trigeminal ganglions, but the sizes of positive neurons were not changed. None of NOS-positive fibers was found in human normal and inflammatory dental pulp tissues. The results suggested that NOS in trigeminal ganglion might play an important role in sensory transmission and regulation of pulpalgia. The absence of NOS-positive nerves in human pulp suggested that NO may not be related to inflammatory stimulation and transmission in dental pulp tissues.

To investigate the expression of interleukin-2 (IL-2), metabotropic glutamate receptor subunit 1 (mGluR1) and estrogen receptor (ER) in neurons of the rat central nervous system (CNS) and identify the coexistence possibility of these immune-neuro-endocrine substances in the central neurons, the tri-labeling immunocytochemical technique with different species-specific primary antibodies (goat anti-IL-2 antibody, rabbit anti-mGluR1 antibody and mouse anti-ER anti-body) were used to incubate two serial neighbor sections (one for demonstrating IL-2, another for mGluR1 and ER) of the cerebral cortex, medulla oblongata and spinal cord. There were IL-2-, mGluR1-and ER-immunoreactivity (IR)-positive labeled neurons in the above-mentioned central areas. The IL-2-IR production showed brown color, located in the cytoplasm; In the neighbor serial section, the mGluR1-IR, production showed blue-black color, located on the cell membranes the ER-IR production also showed brown color, located in the cytoplasm and nuclei. There were mGluR1/ER double-labeled cells in the same section, which accounted for about 50 %–60% of the total single and double labeled neurons. It was identified by projection check of serial neighbor sections that had mGluR1/ ER/IL-2 tri-labeled cells, which accounted for about 30% of total mGluR1/ER double-labeled neurons. The results indicate that mGluR1, ER and Il-2 can coexist in the same rat central neurons, therefore, providing morphological basis for the theory about immune-neuro-endocrine network at the cellular level for the first time.

The inhibition of CH50, a recombinant polypeptide of human fibronectin, on the formation of tumor metastasesin vivo was studied by inoculation of melanoma B16/F1 cells by hypodermic or intraperitoneal injection, and the size and amount of tumor nodes after therapy were measured. In the treatment of hypodermic tumor, local injection of CH50 produced much better efficacy than distance injection of CH50 did. The inhibitory effect of CH50 on the growth of tumor reached 50 %. In the treatment of peritoneal metastasis, the inhibition of CH50 on metastases smaller than 1 mm was above 80 %, and above 50 % on metastases larger than 1 mm. A better efficacy was achieved if CH50 was used in combination with hydroxycamptothecine (HCPT), a chemotherapeutic agent. CH50 displayed a strong inhibitory effect on the formation of small metastasis and growth of larger nodes of tumor, suggesting that CH50 plays a very important role in combined treatment of tumor therapy.