Volume 85  ·   Issue 41  ·  2005

Sepsis means that the inflammatory response caused by pathogenic microorganisms and their products cannot be controlled at an appropriate level, and the response is uncontrolled. The pro-inflammatory cytokines and inflammatory mediators initiate a cascade reaction, and the amplification effect appears, resulting in a violent and abnormal inflammatory reaction, resulting in tissue destruction and organ function damage.

Primary liver cancer (hereinafter referred to as liver cancer) is the second cause of tumor death in China, and its high recurrence rate after radical resection seriously affects the overall surgical efficacy of liver cancer. A large series of clinical data at home and abroad [1,2] show that the recurrence rate of liver cancer after surgery is about 40% ~50% in 3 years, and 60% ~70% or even higher in 5 years. High recurrence rate is also seen in local minimally invasive treatment and after liver transplantation. Exploring the treatment measures of postoperative recurrence of liver cancer is an important topic to effectively prolong the survival time of patients. This exploration mainly focuses on two aspects at present. One is to study the molecular mechanism of recurrence and metastasis of liver cancer. Although important findings have been made so far, there are still many difficulties in forming widely used clinical targeted therapy. The other is to study the role of existing or new comprehensive therapeutic measures in preventing recurrence of liver cancer, which is of practical value to improve the surgical efficacy of liver cancer in the near future.

One of the remarkable characteristics of substance and energy metabolism in severe burn and other pathological states is high catabolism, which can make the body quickly fall into negative nitrogen balance, which seriously affects the prognosis of patients [1-4]. Skeletal muscle, which accounts for about 50% of the total cell weight of the body, is the largest nitrogen pool in human body, and the decomposition and consumption of skeletal muscle protein will undoubtedly have a profound influence on the overall metabolism of the body in severe burn and sepsis. Therefore, it is of great significance to pay further attention to the study of high catabolism of skeletal muscle protein in severe burn and other pathological states [5].

Objective to investigate whether crude extract of Pueraria lobata and puerarin have the same protective effects on the oxidative damage of fetal rat hippocampal cells induced by ethanol. Methods Hippocampal cells were cultured from ICR fetal rats at 18 days of gestation. The changes of HSP70 mRNA and protein expression in fetal rat hippocampal cells induced by ethanol were observed by RT-PCR and Western blot. Results 15 mg/L crude extract of Pueraria lobata and 10 mg/L puerarin can partially antagonize the increase of HSP70 expression in fetal rat hippocampal cells induced by ethanol at 50~350mmol/L concentrations. Conclusion Both crude extract of Pueraria lobata and puerarin can reduce the increase of HSP70 expression in hippocampal cells induced by ethanol, which confirms that both have the same antioxidant effects.

Objective to study the cognitive function and psychological characteristics of chronic fatigue syndrome (CFS) patients in China<Clinical Memory Scale Manual>, SCL-90, Hamilton Depression Scale (HAMD) and Hamilton Anxiety Scale (HAMA) were used to evaluate the memory and psychological tests in 91 CFS patients and 30 healthy controls. Combined with the clinical manifestations, the characteristics of cognitive function and psychological abnormalities in CFS patients were analyzed. Results the memory quotient of CFS group was 85 ± 14, and that of control group was 98 ± 12, t =4.627, P =0.000.The total score of SCL-90 test was 192 ± 47, and that of control group was 140 ± 46, t =5.297, P =0.000; HAMD of CFS patients was 9. 9 ± 6.1, and that of control group was 6.5 ± 2.5, t =2.948, P =0.004; HAMA score of CFS patients was 9.9 ± 7.0, and that of control group was 5.9 ± 2.9, t =3.015, P =0.003. Conclusion The cognitive function of CFS patients in China is significantly impaired, and there are widespread psychological abnormalities, and their characteristics are different from those of primary psychological diseases.

Objective to observe the direct effect of connective tissue growth factor (CTGF) on the transdifferentiation of human renal tubular epithelial cells in vitro, and to investigate the intervention effect of blocking the expression of endogenous CTGF on the transdifferentiation of human renal tubular epithelial cells induced by transforming growth factor-β 1 (TGF-β 1), so as to elucidate the role of CTGF in the transdifferentiation of human renal tubular epithelial cells. Methods Human proximal convoluted tubular epithelial cells (HKC) were cultured in vitro. (2) Low-dose CTGF group: recombinant human CTGF (rhCTGF) was added to the culture medium with a final concentration of 2.5 μ g/L; (3) High-dose CTGF group: the final concentration of rhCTGF was 5.0 μ g/L. To investigate the role of CTGF in the transdifferentiation of HKC induced by TGF-β 1, the cells were divided into four groups: (1) normal control group; (2) TGF-β 1 stimulation group (the culture medium was added with recombinant human TGF-β 1 protein, the concentration was 10.0 μ g/L); (3) TGF-β 1+ CTGF sense oligonucleotide (ODN) group (first transfected with CTGF sense ODN, then stimulated with TGF-β 1); (4) In the TGF-β 1+ CTGF antisense ODN group (first transfected with CTGF antisense ODN and then stimulated with TGF-β 1), the mRNA levels of α-smooth muscle actin (α-SMA) and collagen type IV (col IV) in HKC were determined by reverse transcription-polymerase chain reaction (RT-PCR); Detection of α-SMA expression in cytoplasm of HKC by indirect immunofluorescence method; The percentage of α-SMA positive cells was detected by flow cytometry; The concentration of col Ⅳ in the supernatant of the culture medium was determined by ELISA. Results the α-SMA mRNA levels were significantly increased after different concentrations of rhCTGF were applied to HKC for 24 h (P<0.01), while the expression level of col Ⅳ mRNA decreased significantly (all P<0.01); After 48 h of stimulation, the expression of α-SMA in cytoplasm was significantly enhanced. The positive percentages of α-SMA in the three groups were 2.4%, 38.9% and 65.5% respectively (P<0.01); The results of ELISA showed that rhCTGF inhibited the secretion of col Ⅳ (P<0.01). TGF-β 1 can induce the high expression of CTGF and α-SMA in HKC. 6 h after transfection, CTGF antisense ODN can significantly inhibit the expression of CTGF and α-SMA mRNA in HKC (P<Conclusion CTGF can stimulate the transdifferentiation of renal tubular epithelial cells into myofibroblasts (MyoF) in vitro, and the introduction of CTGF antisense ODN can effectively inhibit the transdifferentiation process induced by TGF-β 1. Therefore, CTGF may be an important factor in regulating the transdifferentiation of renal tubular epithelial cells.

Objective to investigate the role of RASSF1A mRNA expression in cervical cancer tissues and its relationship with HPV-16 infection. Methods RT-PCR was used to detect the expression of RASSF1A mRNA in 56 cervical cancer tissues, 36 cervical intraepithelial neoplasia (CIN) tissues and 20 normal cervical tissues. Results the expression deletion rates of RASSF1A mRNA in normal cervix, CIN and cervical cancer tissues were 0% (0/20), 22.2% (8/36) and 50.0% (28/56), respectively. The expression deletion rate of RASSF1A mRNA in advanced cervical cancer tissues was significantly higher (82.4%) than that in early cervical cancer tissues (35.9%). The expression deletion rate of RASSF1A mRNA in cervical squamous cell carcinoma tissues was significantly higher (63.3%) than that in cervical adenocarcinoma tissues (34.6%) (P<The expression loss rate of RASSF1A mRNA in poorly differentiated cervical cancer tissues (70.8%) was significantly higher than that in moderately and highly differentiated groups (37.5%). The expression loss of RASSF1A mRNA in cervical cancer tissues was negatively correlated with HPV-16 infection (P<0.05), and this relationship is more obvious in cervical adenocarcinoma. Conclusion the expression loss of RASSF1A gene may play an important role in the occurrence and development of cervical cancer.

Objective to construct five eukaryotic expression vectors for expressing shRNA targeting rat kir2.1 gene in tandem and to observe their effects on the expression of this gene.Methods Five RNA interference sites targeting rat kir2.1 gene were selected, and five pairs of oligonucleotide strands were designed and synthesized respectively.After forming double strands, the corresponding vectors with U6 promoter were ligated respectively.The five target genes expressing kir2.1 shRNA were sequentially ligated into the vector pEGFP6-1 by repeated enzyme digestion.The tandem eukaryotic expression vector pEGFP6-1Kir2.1 was constructed by transfecting it into cultured rat cardiomyocytes, and its effects on the transcription of kir2.1 mRNA and protein expression were detected by RT-PCR and Western blot. Results The eukaryotic expression vectors expressing five rat kir2.1 shRNA in tandem were successfully constructed. The inhibition rate of this vector on the transcription of kir2.1 mRNA and the expression of kir2.1 protein in cardiomyocytes was 83.5% and 68.1%

Objective to investigate the effect of carboxy-terminal fragment of osteogenic growth peptide (OGP) [OGP (10~14)] and its derivative G38I on the proliferation and differentiation of rat osteoblasts. Methods Rat osteoblasts of cranial bone were cultured in vitro until the third passage, and 10-15~10-7mol/L of OGP (10~14) or G38I were added respectively. After 48 h, the cells were counted, and the cell proliferation was measured by thiazole blue (MTT) colorimetric assay. The cells were divided into experimental group [OGP (10~14) group and G38I group] and control group. The experimental group was given 10-11mol/L drug intervention for 48 h. The ultrastructural changes of the cells were observed by electron microscope, and the content of alkaline phosphatase (ALP) in the culture supernatant was measured. The changes of type Ⅰ collagen in the cells were compared by immunohistochemistry. The expression levels of nuclear binding factor a1 (Cbfa1) and type Ⅰ collagen mRNA in the cells were measured by reverse transcription-polymerase chain reaction (RT-PCR). Results At lower concentrations, with the increase of drug concentration, the effect of OGP (10~14) and G38I on increasing cell number and increasing cell activity was enhanced. The number of osteoblasts was 37×104/ml and 30×104/ml when the drug was treated by 10-11mol/L, respectively. Further increase of drug concentration inhibited cell proliferation. Under electron microscope, the rough endoplasmic reticulum of the experimental group was well developed, and there were a large number of secretory vesicles in the cytoplasm. Calcium salt crystals were observed in the vesicles of the control group. Compared with the control group, the content of ALP in the culture supernatant of OGP (10~14) and G38I group was higher (4.47 U/g, 3.82 U/g vs 2.21 U/g), and the content of type Ⅰ collagen in the cells was higher<0.01, P<Conclusion OGP (10~14) and its derivative G38I have the same effects of promoting osteoblast proliferation and cell activity as OGP full-length peptide. OGP (10~14) and G38I can up-regulate the mRNA expression of Cbfa1 and type Ⅰ collagen in osteoblasts, increase the synthesis of type Ⅰ collagen in cells, promote cell differentiation and osteogenesis.

Objective to observe the correlation between the expression of signal transducer and activator 3 (Stat3) and cyclooxygenase 2 (COX-2) mRNA in human colorectal malignant tumors and the pathological characteristics of colorectal cancer, and to determine the effect of Stat3 pathway on the proliferation and apoptosis of colorectal cancer cells and the expression of COX-2 gene.Methods The expression of Stat3 and COX-2 mRNA in 350 colorectal cancer tissues and their adjacent normal intestinal mucosa was detected by reverse transcription-polymerase chain reaction (RT-PCR). The expression of p-Stat3 and COX-2 was detected by immunohistochemistry (SP method). AG490, an inhibitor of Stat3 upstream kinase JAK2, was applied to colon cancer cell line HT-29, and the cell proliferation status was detected by tetramethylazolazole blue (MTT) assay; The expression of p-Stat3 and COX-2 protein in colorectal cancer tissues was observed by flow cytometry, and the expression of p-Stat3 and COX-2 protein was detected by Western blot. Results the expression levels of Stat3 and COX-2 mRNA in colorectal cancer tissues were significantly higher than those in normal intestinal mucosa<0.05), 1.97 and 1.88 times of normal tissues, respectively; The expression level of COX-2 was positively correlated with that of Stat3 (correlation coefficient 0.749, P<0.01), and it was related to the degree of tumor differentiation and lymph node metastasis (P<0.05); Immunohistochemical results showed that p-Stat3 was correlated with the expression level of COX-2 protein. Blocking the Stat3 pathway with (AG490) could inhibit the proliferation and induce apoptosis of HT-29 cells; At the same time, the phosphorylation activity of Stat3 decreased with the prolongation of action time, and the expression level of COX-2 also decreased gradually. Conclusion Stat3 pathway has an effect on the proliferation and apoptosis of colorectal cancer cells and the expression of COX-2 gene; Blocking the signal pathway can inhibit the proliferation of colon cancer cells and promote their apoptosis.

Objective to analyze the changes of skeletal muscle protein degradation and its biological mechanism in patients with severe burn sepsis. Methods the blood, 24-h urine and quadriceps muscle samples from 20 patients with severe burn sepsis (burn sepsis group) and 12 patients with plastic surgery (control group) were collected at the same time. The contents of plasma cortisol and tumor necrosis factor (TNF) -α were determined by radioimmunoassay, the contents of 3-methylhistidine (3-MH) and the excretion of 3-MH in urine were detected by high performance liquid chromatography, and the expression of mRNA encoding ubiquitin, E2-14k and proteasome C2 subunit in quadriceps muscle was detected by Western blotting. The expression of proteasome C2 subunit in quadriceps muscle was detected by Western blotting. Results the contents of plasma cortisol (478 ± 56) μ g/L and TNF-α (22.4 ± 3.8) μ g/L in burn sepsis group were significantly higher than those in control group (72 ± 12) μ g/L and TNF-α (0.5 ±<The content of 3-MH in quadriceps femoris (4.3±0.6) μ mol/g and 24-h urinary 3-MH excretion (456±25) μ mol in the burn sepsis group were significantly higher than those in the control group (2.5±0.4) μ mol/g and 24-h urinary 3-MH excretion (202±29) μ mol<The expression of 2.4 kb band and 1.2 kb band of ubiquitin mRNA in quadriceps femoris was increased by 51% and 32%, respectively, compared with the control group, and the difference was statistically significant (P<0.01); The expression of E2-14k mRNA 1.2 kb band increased by 75% compared with the control group, and the difference was statistically significant (P<0.01); The expression of proteasome C2 subunit mRNA and protein increased by 39% and 130%, respectively, compared with the control group (P<Conclusion Protein degradation of skeletal muscle is significantly enhanced in severe burn sepsis, and the content of glucocorticoid and TNF-α in vivo is increased. One of the reasons for enhanced protein degradation in skeletal muscle is the activation of the intracellular protein degradation pathway-ubiquitin system from the gene level in severe burn sepsis.

Objective to observe the protective effect of ulinastatin on several important organ functions after severe burn. Methods Twelve male miniature pigs of Guizhou three lines were randomly divided into two groups: Group A (burn control group, n =6) and Group B (UTI treatment group, n =6), resulting in 35% III degree burn. Among them, 4 animals of Group A and B were randomly selected to draw venous blood as normal controls before burn. From May 2002 to December 2004, 40 patients with 50% ~70% deep Ⅱ ~ Ⅲ degree and Ⅲ degree>30% were randomly divided into conventional treatment group (n =16) and UTI treatment group (n =24). 16 young adult volunteers were the normal control group. Patients in both groups were treated with conventional anti-shock, wound treatment and nutrition therapy. Patients in UTI group were intravenously infused with UTI 400,000 U, twice/d, and patients in conventional group were given equal volume of normal saline for 7 days. The levels of serum LPS, TNF-α, MPO, NE, ALT, AST, CK, CKMB, LDH, BUN and Cr were detected on the 1st, 3rd and 7th days after injury. Results the levels of serum TNF-α, IL-6, D-lactic acid, MDA, ALT, AST, CK, CKMB, LDH, BUN and Cr were significantly increased and the levels of SOD were significantly decreased after severe burn, especially in group A (P<0.05). At 6, 24, 48 and 72 hours after injury, the levels of serum TNF-α (pg/L) in group A were 137 ± 14, 254 ± 10, 202 ± 28 and 183 ± 15, and in group B were 86 ± 15, 177 ± 19, 152 ± 15 and 138 ± 20, respectively. The levels of serum IL-6 (pg/L) in group A were 296 ± 25, 524 ± 42, 422 ± 57 and 373 ± 17, and in group B were 180 ± 32, 388 ± 45, 332 ± 40 and 296 ± 42, respectively. The levels of serum MDA (nmol/ml) in group A were 5.6 ± 0.7, 6.4 ± 0.4, 4.9 ± 0.5 and 4.8 ± 1.0, respectively. Reduce the degree of organ function impairment after burn.

Toxoplasma gondii antigen-IgM (+) was examined in the last menstrual period on December 14, 1997. A routine fetal B-ultrasound examination in June 1998 revealed "space occupying lesions in fetal heart cavity". The diagnosis was: 26 weeks intrauterine pregnancy, 1 pregnancy and 0 delivery, fetal malformation. The patient required midterm induction of labor.

Tumor imaging with radionuclide-labeled antisense oligodeoxynucleotides (ASODN) can detect the tumor tissues that are activated, amplified and overexpressed without forming solid, so as to realize the early diagnosis of tumor [1, 2]. Breast cancer is a major malignant tumor with the highest morbidity and mortality rate in women [3]. If the expression type and clinical stage of oncogene can be identified at the early stage of tumor, and the treatment can be improved by "tailored" treatment. c-erbB2 is a proto-oncogene of breast cancer, which is closely related to malignant transformation and poor prognosis of breast cancer [4]. In this paper, a 15 mer antisense oligodeoxynucleotide complementary to the mRNA of the target gene was designed with c-erbB2 as the target gene, labeled with radionuclide technetium (99mTc), and linked with epidermal growth factor (EGF) to obtain an antisense probe. The probe obtained the introduction characteristics of receptor/ligand specific binding, and passed through the surface of tumor cell membrane This paper aims to study the tissue distribution of antisense probes in tumor-bearing mice and the imaging of tumor tissues with overexpression of oncogene.

There is a subset of T lymphocytes in normal humans and mice, which are named CD4+ CD25+ immunoregulatory T cells (abbreviated as Treg) because they express CD4 and interleukin-2 receptor alpha chain (IL-2R or CD25) surface antigens simultaneously. These immunoregulatory cells are mainly produced in thymus, but can inhibit immune response in peripheral lymphoid organs and maintain the balance of normal immune response and immune tolerance. Treg has been found to play a very important role in preventing and inhibiting autoimmune diseases, inflammation, allergy and transplant rejection. On the other hand, these cells have potential negative effects on tumors, immunodeficiency and viral infections. Therefore, the research on Treg immunoregulatory cells is one of the most active fields in immunology and the whole medical field in recent years, and has a very broad clinical application prospect. The prospect and research progress of Treg in disease occurrence and clinical application are summarized as follows.

Sepsis is a group of clinical manifestations and symptoms of excessive inflammatory response caused by infection. It is a common complication after severe trauma, burn, shock and major surgery, and it is also one of the important causes of death in critically ill surgical patients. At present, it is generally believed that the activation of neutrophil, lymphocyte and monocyte system and the release of endogenous mediators play a key role in the pathophysiological mechanism of posttraumatic sepsis. Among them, the role of cytokine cannot be ignored. According to the different roles of different cytokines in the occurrence and development of sepsis, they can be divided into two categories: pro-inflammatory and anti-inflammatory cytokines.

Matrix metalloproteinase-3 (MMP-3) and matrix metalloproteinase tissue inhibitor-2 (TIMP-2), as important members of MMP and TIMP family, are closely related to the occurrence and development of esophageal carcinoma. We used immunohistochemistry to detect MMP-3 and TIMP-2 in 50 cases of esophageal squamous cell carcinoma (ESC) tissues, and focused on the relationship between them and ESC differentiation degree, lymph node metastasis and other biological behaviors, so as to provide theoretical basis for clinical biomarkers to judge the prognosis of esophageal cancer.

Cervical cancer has long or reversible precancerous lesions before onset, and the main cause of its pathogenesis is human papillomavirus (HPV) infection. However, histopathology and HPV detection cannot evaluate cervical lesions overall, and if one index is found to evaluate the progression or regression of lesions, it will be of great help to further treatment and prognosis of cervical precancerous lesions. cyclin G1 is a new member of cyclins family [1], which is highly expressed in many tissues with active proliferation, and antisense structure can inhibit cell growth [2]. Therefore, it is believed that cyclin G1 is similar to other cyclins or proto-oncogenes, and plays a role in promoting growth. In this experiment, by detecting the expression of cyclin G1 and HPV in different grades of cervical lesions, the relationship between cyclin G1 and high-risk HPV, and the influence on the occurrence and development of cervical lesions were investigated.

Avian influenza virus (H5N1) breaches the boundary between species and causes human infection, so avian influenza is gradually becoming an important disease that threatens human health. In the latest concept column of the New England Journal of Medicine (N Engl J Med, 2005, 353: 1374-1385), published on September 29, 2005, the report of the World Health Organization (WHO) Committee on Works on some contents of the Hanoi Conference on May 10-12, 2005 was published.