A study on the influence of montelukast on chromogranin A protein and mRNA expressions at lung tissue of asthmatic rat / 预防医学

Objective To investigate the potential roles of chromogranin A in pathogenesis of asthmatic inflammation,and to assess the regulation of montelukast on chromogranin A expression.Methods The rat asthma model was established with ovalbumin,and they were allocated to three groups,named asthma group,control group and montelukast group.The expressions of chromogranin A protein and mRNA at lung tissue were detected by immunohistochemisty or real-time PCR methods,and positive expression intensity of chromogranin A protein was assayed by optical density.The correlation between chromogranin A protein and mRNA was also analyzed.Results The expression levels of chromogranin A protein in asthma group(0.34 ±0.05 optical density)was significantly higher than that in control group (0.21 ±0.06 optical density)(P<0.01 ).The expression levels of chromogranin A mRNA in asthma group (4.02 ±0.95 relative quantity value)was significantly higher than that in control group(P<0.01 ).The expression levels of chromogranin A protein in montelukast group(0.28 ±0.04 optical density)was dramatically lower than that in asthma group (0.34 ±0.05 optical density)(P<0.05),while there were no statistical significance of chromogranin A mRNA(3.67 ±0.78 relative quantity value)between those two groups(P>0.05 ).But levels of mRNA was positively correlated with protein of chromogranin A (r=0.635,P<0.01).Conclusion Expressions of chromogranin A protein and mRNA at lung tissue were increased in asthmatic rats,and the results demonstrated that chromogranin A perhaps participated in the pathogenesis of asthma inflammation,but this function of chromogranin A protein could be down regulated by montelukast.

Diagnosis and prognosis of spontaneous pneumomediastinum in eighteen children / 中华儿科杂志

<p><b>OBJECTIVE</b>To analyze the diagnosis, treatment and prognosis of spontaneous pneumomediastinum (SPM) in children.</p><p><b>METHOD</b>A retrospective analysis of the clinical data of 18 children diagnosed with SPM in Yuying Children's Hospital Affiliated to Wenzhou Medical University from December 2007 to February 2013 was performed. Information of the sequelae and recurrence of SPM was obtained by telephone follow-up. SPM was diagnosed according to Versteegh's standard. SPM cases due to mechanical ventilation, trauma, inhaled foreign body or as a result of the underlying disease were not included. Also cases of secondary pneumothorax pneumomediastinum and neonatal mediastinal emphysema were excluded.</p><p><b>RESULT</b>Fifteen of 18 cases were boys and 3 were girls, the range of age was from 9 to 17 years. Predisposing factors included sport activities, severe cough or without a known cause. Clinical manifestations included chest pain, chest tightness, dyspnea, neck pain, back pain, foreign body sensation or pain on swallowing, throat pain of swelling. Chest CT of 18 cases showed pneumomediastinum, 8 cases displayed varied degrees of air in neck, chest; 18 cases of SPM responded well to bed rest, oxygen, antitussive and anti-infection treatment. Fifteen cases received chest CT or X-ray inspection after therapy, showing that the pneumomediastinum disappeared or significantly absorbed, 3 cases improved in clinical symptom. Among 18 patients, telephone follow-up of 14 were successful and 4 cases were lost. An average follow-up time was (24 ± 17) months. None of the cases had any serious consequences, and recurrence happened in one case.</p><p><b>CONCLUSION</b>Children's spontaneous pneumomediastinum is a benign disease. When a child has chest pain or chest tightness, SPM should be considered after excluding the common diseases. SPM can be diagnosed in association with clinical feature and chest CT examination. Patients respond well to conservative therapy and most of them had no severe sequelae.</p>

Effect of budesonide on the expression of urotensin-II and its mRNA in asthma airway remodeling rats / 中国药学杂志

OBJECTIVE: To study the effects of budesonide(BUD) on the expression of urotensin-II (UII) and its mRNA in asthma airway remodeling rats. METHODS: Twenty-four Sprague-Dawley (SD) rats were randomly divided into three groups: the control group, asthma group and BUD treated group. In the experiment, the models of asthma were established by sensitization and challenge with ovalbumin (OVA). The total brochial wall thickness (Wat) and the airway smooth musle thickness (Wam) were measured by image analysis system. The levels of UII in BALF and serum were measured by enzyme linked immunosorbent assay (ELISA). The protein and mRNA expression of UII were detected by immunohistochemistry and reverse transcription polymerase chain reaction (RT-PCR), respectively. RESULTS: Wat and Warn of of asthma group were significantly higher than those of the control group (P < 0.01, respectively). Compared with asthma group, those were all markedly decreased in BUD group (P < 0.01, respectively). The levels of UII in BALF and serum of asthma group were markedly higher than those in control group and BUD group (P < 0.01, respectively). Immunohistochemistry and RT-PCR showed that the protein and mRNA expression of UII in asthma group were significantly higher than those of control group (P < 0.01, respectively). While those of BUD group were significantly lower than those of asthma group(P < 0.01, respectively). There were a significantly positive correlation between Wat and the protein and mRNA expression of UII. At the same time, there were a significantly positive correlation between Warn and the protein and mRNA expression of UII. CONCLUSION: The beneficial effect of glucocorticoid (BUD) on asthma airway remodeling may be at least in part due to their direct inhibitory effect on UII. Copyright 2012 by the Chinese Pharmaceutical Association.

Effects of dexamethasone on 15-lipoxygenase expression in lungs of asthmatic rats / 中国当代儿科杂志

<p><b>OBJECTIVE</b>15-lipoxygenase (15-LO) is a prooxidant enzyme which is expressed in asthmatic lungs leading to formation of pro- and anti-inflammatory mediators. Gene expression profiling studies show the association between 15-LO and allergic asthma. This study was designed to observe the expression of 15-LO in lungs of asthmatic rats and examine the effects of dexamethasone on 15-lipoxygenase expression.</p><p><b>METHODS</b>Twenty-seven male Sprague-Dawley (SD) rats were randomly divided into three groups: control, asthma and dexamethasone (DXM) intervention. An asthma model was prepared by sensitization and challenging with ovalbumin. The production of 15-LO in lung tissue homogenates was measured using ELISA.The expression of 15-LO mRNA in lungs was determined by reverse transcription-polymerase chain reaction (RT-PCR).</p><p><b>RESULTS</b>The levels of 15-LO mRNA and protein in the asthma group (0.51 ± 0.14 and 2080 ± 73 μg/mL, respectively) were lower than those in the control group (0.76 ± 0.15 and 2472 ± 106 μg/mL, respectively; P<0.01). DXM intervention increased significantly the levels of 15-LO mRNA and protein (1.02 ± 0.34 and 2562 ± 218 μg/mL) compared with the asthma group (P<0.01).</p><p><b>CONCLUSIONS</b>The production of 15-LO in lung tissues is reduced in asthmatic rats. DXM can increase the expression of 15-LO in lung tissues and thus might provide anti-inflammatory effects in asthmatic rats.</p>

Effects of lentinan on interleukin-1β-induced transdifferentiation of human embryonic lung fibroblasts to myofibroblasts / 中国当代儿科杂志

<p><b>OBJECTIVE</b>To study the effects of interleukin-1β (IL-1β) on transdifferentiation of human embryonic lung fibroblasts to myofibroblasts and the effects of lentinan on the transdifferentiation.</p><p><b>METHODS</b>The human embryonic lung fibroblasts were cultured in vitro, and fibroblasts were treated with different concentrations of IL-1β and lentinan. The proliferation activity of the human embryonic lung fibroblasts was evaluated by the Cell Counting Kit-8 (CCK-8). The expression of α-smooth muscle actin (α-SMA) protein was measured by immunocytochemistry. The levels of fibronectin (FN), typeⅠcollagen (ColⅠ) and α-SMA mRNA were detected by RT-PCR.</p><p><b>RESULTS</b>Compared with the untreated control group, the absorbance value of cell proliferation, α-SMA protein levels, FN, ColⅠand α-SMA mRNA expression were significantly up-regulated after different concentrations of IL-1β (0.1, 1, 10 ng/mL) treatment for 48 hrs (P<0.01). Lentinan treatment inhibited up-regulation of the cell proliferation activity, α-SMA protein levels, FN, ColⅠand α-SMA mRNA expression induced by IL-1β in a dose-independent manner (P<0.01).</p><p><b>CONCLUSIONS</b>Lentinan can suppress human embryonic lung fibroblast proliferation, fibroblast-myofibroblast transdifferentiation and extra cellular matrix synthesis induced by IL-1β.</p>

Expression of Rock2 and TGF-β1 mRNA and glucocorticoid intervention in asthmatic rats / 中国当代儿科杂志

<p><b>OBJECTIVE</b>To study the roles of rhoassociated coiled coil forming protein kinase 2 (Rock2) and transforming growth factor-β1 (TGF-β1) mRNA in acute asthma and the effect of glucocorticoid intervention on the Rock2 and TGF-β1 mRNA expression in rats.</p><p><b>METHODS</b>Forty-eight male rats were randomly divided into 4 groups (n=12 each): asthma, control, dexamethasone treated (DXM) and budesonide treated (BUD). Rat model of asthma was prepared by the ovalbumin (OVA) challenge. The animals were sacrificed 24 hrs after the last challenge. The total cell number and differentiation cell number were counted in bronchoalveolar lavage fluid (BALF). The protein expression of Rock2 was ascertained by immunohistochemistry and the mRNA expression of TGF-β1 was ascertained by hybridization in situ.</p><p><b>RESULTS</b>The pathological changes in the BUD and the DXM groups were alleviated when compared with the asthma group. The total cell number and the percentage of eosinophil (EOS), polymorphonuclear leukocytes (PMN) and lymphocytes (Lym) in BALF in the asthma group were significantly higher than those in the control group (P<0.01). The percentage of macrophage (Mф) in the asthma group was significantly lower than that in the control group (P<0.01). The total cell number and the percentage of EOS and Lym in BALF in the DXM and the BUD groups decreased, while the percentage of Mф increased significantly compared with those in the asthma group (P<0.01). The Rock2 and TGF-β1 mRNA expression in lung tissues in the asthma group increased significantly compared with those in the control, BUD and DXM groups, while there were no significant differences in the Rock2 expression and TGF-β1 mRNA expression between the DXM or BUD group and the control group.</p><p><b>CONCLUSIONS</b>The expression of Rock2 and TGF-β1 mRNA in lung tissues is increased in rats with acute asthma. Glucocorticoids can significantly decrease the expression of Rock2 and TGF-β1 in lung tissues, thus alleviates airway inflammation.</p>

Changes in urotensin-II expression in airway remodelling in asthmatic rats / 中国当代儿科杂志

<p><b>OBJECTIVE</b>To study the role of urotension-II in serum and bronchoalveolar lavage fluid (BALF) in the process of airway remodelling in asthmatic rats.</p><p><b>METHODS</b>Thirty-two male Sprague-Dawley (SD) rats were randomly divided into normal control and 2-week, 4-week and 8-week asthmatic groups (OVA inhalation of 2, 4 and 8 weeks respectively). Rats were sensitized and challenged by OVA to establish a model of asthma. The bronchial wall thickness and the airway smooth muscle thickness were measured by image analysis system. The urotension-II contents in serum and BALF were determined using ELISA.</p><p><b>RESULTS</b>The bronchial wall thickness and the airway smooth muscle thickness in the three asthmatic groups significantly increased compared with those in the normal control group (P<0.01). The urotension-II contents in serum and BALF in the three asthmatic groups also increased significantly compared with those in the normal control group (P<0.01). The urotension-II contents in serum and BALF in the 8-week asthmatic group were the highest, followed by the 4-week and the 2-week asthmatic groups (P<0.01). BALF urotension-II contents were positively correlated with the bronchial wall thickness and the airway smooth muscle thickness as well as serum U-II contents in the four groups.</p><p><b>CONCLUSIONS</b>The urotension-II contents in serum and BALF in the process of airway remodeling increase in asthmatic rats. The changes in serum and BALF urotension-II contents may be associated with airway remodeling in asthmatic rats.</p>

Clinical study of strengthening pi and nourishing shen therapy combined with Western medicine on patients with glucocorticoid resistant myasthenia gravis / 中国中西医结合杂志

<p><b>OBJECTIVE</b>To study the effect of strengthening Pi and nourishing Shen therapy (SPNST) in treating patients with glucocorticoid resistant myasthenia gravis (GR-MG).</p><p><b>METHODS</b>Twenty-seven patients with MG were enrolled, who were relapse cases after treated by cholinesterase inhibitor with systemic glucocorticoid treatment and showed resistance to glucocorticoid. All were treated by Western medicines, methylprednisolone (MP) and pyridostigmine bromide (PSB), together with Chinese medicine (CM) given according to their syndrome types, namely, for the 15 patients of Pi-Shen qi-yin deficiency type, Buzhong Yiqi Pill and Liuwei Dihuang Pill, and for the 12 patients of Pi-Shen yang-deficiency type, Buzhong Yiqi Pill and Zishen Yutai Pill. The dosages of medicines were reduced gradually in MP-PSB-CM order along with the progressing of the therapy in 4 stages (symptom curing, choline receptor restoration, immune regulation, and functional strengthening). Muscle strength and overall state of patients were re-examined before and after each of the 4 stages.</p><p><b>RESULTS</b>After 1-year treatment, the therapeutic effect in 9 patients was judged as completely remitted; in 7 as remitted with continuous medication; in 5, 1 and 3 as significantly improved, moderately improved and unchanged respectively, while 2 patients died.</p><p><b>CONCLUSION</b>Integrative medicine shows definite effects in treating GR-MG, and it is worthy of further studying.</p>

Clinical analysis of 234 cases with congenital malformations of respiratory system / 中华儿科杂志

<p><b>OBJECTIVE</b>To explore clinical characteristics, radiographic findings and diagnostic methods of patients with congenital malformations of respiratory system for enhancing the diagnosis of congenital malformations of respiratory system in children.</p><p><b>METHOD</b>Totally 234 patients with congenital malformations of respiratory system were chosen from the inpatient department of Yuying Children's Hospital Affiliated to Wenzhou Medical College from July 2003 to June 2008. The clinical presentations and radiographic findings of these children were analyzed.</p><p><b>RESULT</b>Of the 234 patients with congenital malformations of respiratory system, the age at diagnosis was between the first day and 14 years of age, mean age was 1.12 years. The main symptoms were persistent laryngeal stridor, recurrent wheezing, recurrent respiratory tract infections and dyspnea. Through the use of chest X-ray, spiral CT 3D reconstructions, fiberoptic bronchoscopy and other laboratory techniques, 213 cases were diagnosed as having single malformation and 21 cases were found to have multiple malformations. Of the 213 cases with single malformation, 97 cases had laryngeal malformation (congenital laryngeal stridor in 90 cases, congenital laryngeal webs in 5 cases and congenital laryngeal cyst in 2 cases), 35 cases had tracheal-bronchial malformation (congenital tracheobronchial stenosis in 17 cases, congenital abnormal bronchial origin in 7 cases, tracheobronchomalacia in 10 cases and tracheoesophageal fistula in 1 case), 43 cases had lung malformation (pulmonary sequestration in 5 cases, congenital lung cysts in 22 cases, congenital lobar emphysema in 1 case, agenesis of lung and hypoplasia of lung in 8 cases and congenital cystic adenomatoid malformation in 7 cases), 38 cases had diaphragm malformation, 28 cases had congenital tracheal-bronchial stenosis as confirmed by spiral CT 3D reconstructions and fiberoptic bronchoscopy. Ten cases with congenital abnormal bronchial origin were diagnosed with spiral CT 3D reconstructions. Laryngeal stridor and tracheobronchomalacia were diagnosed by fiberoptic laryngoscope and fiberoptic bronchoscopy. The accuracy rates of preoperative diagnosis through clinical and radiographic examinations of 37 cases with lung malformation and 36 cases with diaphragm malformation were 83.78% and 91.67%.</p><p><b>CONCLUSION</b>Congenital malformations of respiratory system are a group of diseases that are important for pediatric respiratory clinicians. Congenital malformations of respiratory system should be considered in children with persistent laryngeal stridor, recurrent wheezing, recurrent respiratory tract infections and dyspnea. The radiographic examination and respiratory endoscope play important roles in the diagnosis of congenital malformations of respiratory system.</p>

Role of c-Jun N-terminal kinase signal transduction pathway in the course of airway remodeling of asthma rat / 中华儿科杂志

<p><b>OBJECTIVE</b>To study the role of c-Jun N-terminal kinase (JNK) signal transduction pathway in the course of asthma airway remodeling, to explore whether IL-1beta participates in asthma airway remodeling mediated by JNK signal transduction pathway.</p><p><b>METHODS</b>Totally 72 male Sprague-Dawlay rats (6 - 8 weeks old, weighing about 120 g) were randomly divided into control groups (36 rats) and asthma groups (36 rats). The rats were sensitized for inducing asthma by intraperitoneal injection of ovalbumin and AL(OH)3 and were repeatedly exposed to aerosolized ovalbumin for 4, 8, 12 weeks (A4, A8, or A12 group), each had 12 rats, and correspondingly control rats were intraperitoneally injected with 0.9% NaCl, then were repeatedly exposed to 0.9% NaCl for 4, 8, 12 weeks (C4, C8, or C12 group), each had 12 rats. The ultrastructural changes of pulmonary tissues were observed by transmission electron microscope (TEM). The total bronchial wall thickness (Wat) and the airway smooth muscle thickness (Wam) were measured by an image analysis system. The concentrations of IL-1beta in serum and bronchoalveolar lavage fluid (BALF) were tested by a "sandwich" ELISA. The protein expressions of P-JNK and P-c-Jun were detected by immunohistochemical technique. Lung tissue extracts were analyzed for phosphorylation of JNK by Western blotting. Linear correlation analysis showed the correlation between Wat and P-JNK protein, Wam and P-JNK protein, levels of IL-1beta in serum and P-JNK protein, levels of IL-1beta in BALF and P-JNK protein.</p><p><b>RESULTS</b>In asthma groups, TEM showed alveolar septal proliferation and alveolus type II epithelial cells swelling. Wat and Wam in all asthma groups were significantly higher than those in corresponding control groups (P < 0.01, respectively), and compared with group A4 and group A8, Wat and Wam of group A12 significantly increased (P < 0.01). The concentrations of IL-1beta in serum and BALF of asthma groups were all significantly higher than those of the corresponding control groups (P < 0.01, respectively), and compared with group A4 and group A8, the concentrations of IL-1beta in BALF of group A12 significantly increased (P < 0.01 or P < 0.05), but the levels of IL-1beta in serum were not significantly different among them (P > 0.05). Mean absorbance values (by immunohistochemistry) of P-JNK and P-c-Jun in asthma groups were significantly higher than those in corresponding control groups (P < 0.01, respectively), and compared with group A4 and group A8, those of group A12 significantly increased (P < 0.01 or P < 0.05). The absorbance (by Western Blot) of P-JNK in A4, A8, A12 group was significantly higher than that in C4, C8, C12 groups (P < 0.01, respectively), and compared with group A4, that of P-JNK of A12 significantly increased (P < 0.01), and compared with group A8, there was no significant difference (P > 0.05). Strong positive correlations were found between Wat or Wam and P-JNK (r = 0.823 and r = 0.818, P < 0.01, respectively, n = 68) and between P-JNK and concentration of IL-1beta in serum or BALF (r = 0.717 and r = 0.803, P < 0.01, respectively, n = 68).</p><p><b>CONCLUSIONS</b>The expression of P-JNK and its downstream P-c-Jun in rats of asthma airway remodeling is increased, which implicates that JNK signal transduction pathway plays an important role in the course of asthma airway remodeling. IL-1beta participates in asthma airway remodeling possibly partly through activating JNK signal transduction pathway.</p>

Adenosine deaminase deficiency associated severe combined immunodeficiency with disseminated varicella infection after vaccination: a case report / 中华儿科杂志

<p><b>OBJECTIVE</b>To enhance the knowledge of adenosine deaminase (ADA) deficiency associated severe combined immunodeficiency (SCID) with disseminated varicella infection after vaccination.</p><p><b>METHODS</b>With case report and review of literature, the background knowledge, clinical and laboratory findings, diagnosis and treatment of ADA-deficient SCID were discussed.</p><p><b>RESULTS</b>The patient had the condition with failure to thrive. The main complaint was more than three weeks of fever and rash. He had received the live attenuated Oka strain varicella vaccination approximately two weeks before the onset of rash. Varicella infection was confirmed with direct immunofluorescence assay. The patient had mild leukopenia, with 3% lymphocytes. The initial immunologic workup included decreased IgG, IgM and IgA, abnormal expanded lymphocyte enumeration which confirmed the reduction of CD3, CD4, CD8, CD19 and CD56. Enzyme testing for ADA activity showed remarkably low level in the hemolysate, as well as increased levels of deoxyadenosine nucleotides.</p><p><b>CONCLUSION</b>ADA-deficient SCID has some characteristic clinical and laboratory findings. Management options for ADA-deficient SCID include hematopoietic stem cell transplantation, enzyme replacement therapy and gene therapy. Immunodeficiency should be considered in children with severe failure-to-thrive. Live vaccine administration should be avoided in patients with immunodeficiency.</p>

Role of external signal regulated kinase signal transduction pathway in airway remodeling of rats with asthma and regulation by glucocorticoids / 中华儿科杂志

<p><b>OBJECTIVE</b>Airway remodeling in asthma makes treatment of asthma very difficult, and study of its pathogenesis becomes very important. The present study aimed to explore the role of external signal regulated kinase (ERK) signal transduction pathway in airway remodeling in rats asthma model and regulatory effects of glucocorticoids on ERK signal transduction pathway and airway remodeling.</p><p><b>METHODS</b>Totally 80 male Sprague-Dawlay rats (6-8 weeks old, weighing about 120 g) were randomly divided into control groups (30 rats), asthma groups (30 rats) and treated groups [including a group intervened with dexamethasone (DM group) and budesonide (BUD group), each had 10 rats]. The rats were sensitized for inducing asthma by intraperitoneal injection of ovalbumin and Al (OH)(3) and were repeatedly exposed to aerosolized ovalbumin for 4, 8, or 12 weeks [respectively called 4, 8 or 12 wk asthma group (A4, A8 or A12 group), each had 10 rats]; and correspondingly control rats were intraperitoneally injected with 0.9% NaCl, then were repeatedly exposed to 0.9% NaCl for 4, 8, or 12 weeks [respectively called 4, 8 or 12 wk control group (C4, C8 or C12 group), each had 10 rats]; DM group rats were repeatedly exposed to aerosolized ovalbumin for 8 wk, and BUD group rats for 12 wk. Total bronchial wall thickness (Wat) and smooth muscle thickness (Wam) were measured by an image analysis system. Concentrations of PDGF-AB in serum were measured by sandwich ELISA. Phospho-ERK (P-ERK) and c-Fos were detected by immunohistochemical technique; lung tissue extracts were analyzed for phosphorylation of ERK by Western blotting.</p><p><b>RESULTS</b>Wat and Wam in all asthma groups were significantly higher than those in corresponding control groups (P < 0.01, respectively), those of the treated groups were significantly lower than asthma groups (P < 0.01). The concentrations of PDGF-AB in serum of asthma groups [(228 +/- 18) pg/ml, (293 +/- 77) pg/ml, (225 +/- 66) pg/ml for A4, A8, A12 groups, respectively] were all significantly higher than those of the control groups [(160 +/- 14) pg/ml, (165 +/- 29) pg/ml and (164 +/- 27) pg/ml for C4, C8, C12 group, respectively] (P < 0.01 or P < 0.05); the value of DM group [(157 +/- 46) pg/ml] was significantly lower than that of the group A8 (P < 0.01), no significant difference was found when the values of BUD group [(208 +/- 40) pg/ml] was compared with that of A12 group (P > 0.05). Mean absorbance values (by immunohistochemistry) of P-ERK and c-Fos in asthma groups were significantly higher than those in corresponding control groups (P < 0.01, respectively), DM group had a significantly lower value than group A8 (P < 0.01), BUD group had a significantly lower value than group A12 (P < 0.01); absorbance (by Western blot) of P-ERK in A4, A8, A12 group was significantly higher than that in C4 and C8 group, the value of DM group was significantly lower than that of group A8 (P < 0.01), and that of BUD group (1.8 +/- 0.2) was significantly lower than that of group A12 (P < 0.01).</p><p><b>CONCLUSION</b>Asthmatic rats have higher concentrations of PDGF-AB in serum and phosphorylation of ERK and c-Fos; glucocorticoids inhibit phosphorylation of ERK and c-Fos in asthmatic rats, and to some extent also inhibit Wat and Wam.</p>

Carrier genetic diagnosis of intron and/or exon-deletion Duchenne muscular dystrophy by microsatellite analysis and quantitative polymerase chain reaction / 中华医学遗传学杂志

<p><b>OBJECTIVE</b>To detect the female carriers from the intron and/or exon-deletion Duchenne/Becker musclular dystrophy (DMD) familial members for prenatal or preimplantation genetic diagnosis.</p><p><b>METHODS</b>Using method of PCR to five microsatellite markers (located in 5' terminus and intron 44, 45, 49, 50), analysing of the short tandem repeat sequence polymorphism with the genescan and binding with the quantitative polymerase chain reaction, we detected the DMD carriers from 1 intron and exon -deletion family and 1 intron-deletion family.</p><p><b>RESULTS</b>The STR-50 genotype of II 2 in family 5 was 245/245, so II3 is DMD gene carrier. The STR-45 genotype of II6 and II8 were del/172, III19 was del/178, so they were all DMD gene carriers.</p><p><b>CONCLUSION</b>The STR haploid linkage analysis combined with quantitative polymerase chain reaction is accurate and efficient to detect the female carriers from the intron and/or exon-deletion DMD familial members.</p>

Regulative mechanism of dexamethasone on Toll-like receptor 4 signal transduction of infant asthma rat / 中华儿科杂志

<p><b>OBJECTIVE</b>Eosinophilic airway inflammation is one of the basic characteristics of allergic asthma. Toll-like receptor is one of the most important innate immunity pattern recognition receptors. Glucocorticoids (GCS) are still the most effective treatment for asthma. However, few reports of studies on regulatory mechanism of GCS on the innate immunity system are available. The mechanism of effects of GCS on TLR4 is unclear. The present study aimed at understanding the effect of dexamethasone (DXM) on change of TLR4 and mechanism of regulatory effect of TLR4 on eosinophil (EOS) apoptosis.</p><p><b>METHODS</b>Twenty-seven Sprague-Dawley (SD) rats (age 28 to 42 days, body weight 120 to 180 gram) were randomly divided into the control group, asthma group and DXM group with 9 in each. Asthma model rats were sensitized with the mixture of ovalbumin (OVA, 1 mg) and Al (OH)(3), 100 mg on day 1 and day 8, repeatedly exposed to aerosolized OVA after day 15, once a day for three days and continued for 30 minutes at every time. During the sensitization stage, 100 microg/ml DXM were prepared with DXM group for every other day, and the same doses DXM were prepared for every day on the stage of challenge. The histopathological changes of lung tissues were observed with light microscope (LM). EOS and other inflammatory cells in bronchoalveolar lavage fluid (BALF) were counted; the concentrations of OVA-sIgE in serum were measured by using "sandwich" ELISA; The expressions of TLR4 mRNA were determined by in situ hybridization, the apoptosis of EOS was detected by TUNEL.</p><p><b>RESULTS</b>(1) LM showed many inflammatory cells infiltration around the bronchi and blood vessels, bronchus mucus increased, airway epithelium damage and desquamation, and airway mucous plugs in asthma group, whereas DXM group showed significantly milder changes. (2) Inflammationary cells count in BALF of asthma group was significantly higher as compared to control group (P < 0.01); compared with asthma group, the total cell count, EOS absolute count and EOS% were all significantly decreased in DXM group [(2.14 +/- 0.10) x 10(9)/L, (4.78 +/- 1.23) x 10(7)/L, (2.17 +/- 0.25)%]. (3) Levels of OVA-sIgE in serum of asthma group [(83.40 +/- 6.80) microg/ml] were significantly higher than those of the control group [(14.38 +/- 4.25) microg/ml] (P < 0.01), while those of DXM group [(45.02 +/- 7.47) microg/ml] were significantly lower than asthma group (P < 0.0 1). (4) There were no significant differences in TLR4 mRNA detected by in situ hybridization between control group (24.71 +/- 0.85) and asthma group (25.81 +/- 3.56) (P > 0.05); but it significantly increased in DXM group (29.86 +/- 3.92) as compared to asthma group. (5) The percentages of apoptotic EOS in asthma group [(7.39 +/- 1.93)%] were significantly lower than those in control group [(9.06 +/- 1.52)%] (P < 0.01); and significantly higher in DXM group [(13.33 +/- 1.09)%] than in asthma group (P < 0.01). There were significantly positive correlations between TLR4 mRNA and the percentage of apoptotic EOS (r = 0.612, P < 0.01).</p><p><b>CONCLUSION</b>DXM can decrease OVA-sIgE level, induce EOS apoptosis, which may correlate with the activation of TLR4 signal transduction.</p>

Effect of achyranthes bidentata polysaccharides on signal transducer and activator of transcription 6 and its mRNA expression in a rat model of asthma / 中国应用生理学杂志

<p><b>AIM</b>To study the effect of achyranthes bidentata polysaccharides(ABPS) on the expression of signal transducer and activator of transcription 6 and its mRNA in bronchus of a rat model of asthma.</p><p><b>METHODS</b>Thirty male SD rats were randomly divided into three groups: the control group, asthma group and ABPS group. The total cell numbers, eosinophils (EOS) numbers and differentiated cell numbers in bronchoalveolar lavage fluid (BALF) were counted by different count fluids. The concentrations of IL-4 in serum and BALF were measured by sandwich ELISA. The protein expressions of STAT6 were detected by immunohistochemistry techniques. The mRNA expressions of STAT6 were detected by hybridization in situ.</p><p><b>RESULTS</b>(1) The total cell numbers in BALF, the absolute numbers of EOS, the ratios of eosinophils to the total cell numbers (EOS%) of asthma group were all significantly higher than those of the control group (P < 0.01). The total cell numbers in BALF, the absolute numbers of EOS and EOS% of ABPS group were all significantly lower than those of asthma group (P < 0.01). (2) The concentrations of IL-4 in BALF and serum of asthma group were significantly higher than those of control group (P < 0.01), while the concentrations of IL-4 in BALF and serum of ABPS group were significantly lower than those of asthma group. (3) Immunohistochemistry showed that the protein content of STAT6 around the bronchus of asthma group was significantly higher than that of the control group (P < 0.01), while that of ABPS group was significantly lower than that of asthma group , the epithelial cells were the chief expression cells; hybridization in situ showed that the mRNA expression of STAT6 around the bronchus of asthma group was significantly higher than that of the control group (P < 0.01), while that of ABPS group was significantly lower than that of asthma group , the epithelial cells were the chief expression cells.</p><p><b>CONCLUSION</b>STAT6 protein and STAT6 mRNA were found strongly expressed in rat asthma model, the epithelial cells were the chief expression cells. ABPS had an inhibitory effect on airway inflammation cells infiltration such as EOS, it significantly depressed STAT6 and its mRNA expression, thus reduced the synthesis of IL-4 might be key in modulating mechanism of asthma.</p>

Therapy of Duchenne muscular dystrophy with umbilical cord blood stem cell transplantation / 中华医学遗传学杂志

<p><b>OBJECTIVE</b>To analyze a Duchenne muscular dystrophy(DMD) patient's muscular regeneration, dystrophin expression and locomotive variation before and after he underwent umbilical cord blood stem cell transplantation in order to assess the therapeutic effect.</p><p><b>METHODS</b>A 12-year-old DMD boy who could not walk for 3 years was confirmed by gene analysis and dystrophin protein immune test on his muscle. He had no other chronic disease. By HLA matching, a piece of umbilical cord blood stem cell with 6 HLA sites matching to the boy was found in Guangdong Umbilical Cord Blood Bank. The number of the nucleated cells of the umbilical cord blood stem cell was 24.08x 10(8). After pretreatment for the DMD boy with busulfan, cyclophosphamide and rabbit anti-human thymocyte globulin, the allergenic cord blood stem cells were transplanted into him by intravenous injection. Cyclosporin A, methylprednisolone, MMF, prostaglandin E1 and ganciclovir were given after the transplantation. At the same time, Gran, the granulocytic cell stimulating factor, and gamma globulin were administered. The biochemistry profile including serum creatine kinase (CK), the reconstruction of blood making, the deletion exon of DMD gene, the regenerating muscles, the dystrophin protein expression, and the locomotive function of the DMD boy were tested regularly.</p><p><b>RESULTS</b>(1) The white blood cells (WBC) of peripheral blood decreased gradually to zero after pretreatment. In a period of 15 days after transplantation, the neutrophil increased to 0.5x 10(9)/L; at 25 days, WBC increased to normal level. Blood platelet was more than 20x 10(9)/L at 22 days. The hemoglobin rose to 85-100 g/L. At 140 days, sternal puncture revealed the rapid growth of neutrophil, blood platelet and hemoglobin. (2)At 140 days, the blood type of the DMD boy transformed from type O to type AB (the donor's blood type being AB). There was no grafe versus host reaction. (3) At 18, 30, 43, 55, 74 and 233 days after transplantation, the PCR-short tandem repeat test of the boy's peripheral blood DNA showed that his genotype was completely the same as the donor's. The results of PCR-short tandem repeat tests of the bone marrow cells DNA by sternal puncture at 140, 183 and 235 days were the same as those of the blood DNA. (4) At 60 days, DMD gene analysis by PCR showed that the defected DMD gene (exon 19 deletion) had been corrected by the umbilical cord stem cells transplantation. (5) At 75 days, the biopsy of calf muscle showed there were myoblast cells and muscular tubes growing. The dystrophin expressions were weak, but a few of them were strong. DNA analysis showed that the donor's gene DNA accounted for 1%-13%. At 126 days, obviously increased dystrophin positive muscular fibers of the boy were found. The donor's fibers rose to 2.5%-25%. (6) The serum CK of the boy declined from 5735 U/L to 274 U/L. (7) At 100 days, physical examination revealed improvement in his arms and legs.</p><p><b>CONCLUSION</b>The therapy of Duchenne muscular dystrophy with allogeneic umbilical cord blood hematopoietic stem cell transplantation may reset up the blood-making function, decrease the serum CK level, restore the dystrophin in muscles, and improve the locomotive function of the DMD boy. These data suggest that the allogeneic umbilical cord blood hematopoietic stem cell transplantation may benefit the DMD boys.</p>

Carrier detection of Duchenne/Becker muscular dystrophy in Chinese families by microsatellite analysis / 中华医学遗传学杂志

<p><b>OBJECTIVE</b>To screen and detect the female carriers from the DMD/BMD family members for prenatal or preimplantation genetic diagnosis.</p><p><b>METHODS</b>For the detection of DMD/BMD carriers from 27 family members in 4 families, PCR to five microsatellite markers(located in 5' terminus and intron 44, 45, 49, 50) and analysis of the short tandem repeat(STR) sequence polymorphism with the use of genescan were implemented.</p><p><b>RESULTS</b>Six of the 17 female members were obligate DMD gene carriers according to the haplotype analysis of the results of the genescan, which conformed with the pedigree analysis. Besides, the authors detected five carriers and five normal females in these families with the use of the haplotype analysis only. The most polymorphic locus was STR 49, and the least was STR 50.</p><p><b>CONCLUSION</b>The STR haploid linkage analysis using (CA)n repeats within the human dystrophin gene is a rapid,accurate, objective method and is well suited for routine use in clinical laboratories engaged in DMD/BMD linkage analysis for the detection of carrier.</p>

Single cell analysis of some deletion in dystrophin gene exons and gender determination by 3-plex nested PCR / 中华医学遗传学杂志

<p><b>OBJECTIVE</b>To set up a technique of single lymphocytes 3-plex nested PCR for dystrophin and SRY gene, and to evaluate the possibility of using this technique for preimplantation genetic diagnosis(PGD) of deleted Duchenne muscular dystrophy (DMD) with family history.</p><p><b>METHODS</b>Fifty single lymphocytes of a normal male and fifty of a normal female were obtained for detecting dystrophin gene(exon 51, exon 19, exon 48) and SRY gene by 3-plex nested PCR.</p><p><b>RESULTS</b>In the group of exon 51/exon 19/SRY, the amplification rates of exon 51, exon 19 and SRY in male were 96%, 94% and 94%; the amplification rates of exon 51 and exon19 in female were 94% and 94%, respectively. In the exon 48/exon 19/SRY group, the amplification rates of exon 48, exon 19 and SRY in male were 92%, 90% and 94%, the amplification rates of exon 48, exon 19 in female were 94% and 92%, respectively.</p><p><b>CONCLUSION</b>The technique of single lymphocytes 3-plex nested PCR for dystrophin and SRY gene established in this study is highly sensitive, specific and reliable, and is suitable for PGD of deleted DMD with family history.</p>

Effect of macrophage inflammatory protein-1alpha and its mRNA on airway inflammation of mouse asthma model / 中华儿科杂志

<p><b>OBJECTIVE</b>To study the effect of macrophage inflammatory protein-1alpha(MIP-1alpha) and its mRNA on airway inflammation of mouse with induced asthma.</p><p><b>METHODS</b>Seventy male BALB/C mice were randomly divided into the control group and asthma group (including 7 subgroups, 10 mice each). The control group included group A(24) (the lavaging subgroup was sacrificed 24 h after the last challenge) and group A(0) (the non-lavaging subgroup was sacrificed from 18 h to 24 h after the last challenge); asthma group included group B(3) (the lavaging subgroup was sacrificed 3 h after the last challenge), group B(8) (the lavaging subgroup was sacrificed 8 h after the last challenge), group B(24) (the lavaging subgroup was sacrificed 24 h after the last challenge), group B(36) (the lavaging subgroup was sacrificed 36 h after the last challenge) and group B(0) (the non-lavaging subgroup was sacrificed from 18 h to 24 h after the last challenge). In the experiment, the mice model of asthma was established by the ovalbumin (OVA) challenge methods. Eosinophils (EOS) numbers and differentiated cell numbers in bronchoalveolar lavage fluid (BALF) were counted; the concentrations of MIP-1alpha in serum and BALF were measured by sandwich enzyme-linked immunosorbent assay (sandwich ELISA); the protein expressions of MIP-1alpha were detected by immunohistochemical techniques; the mRNA expressions of MIP-1alpha were determined by in situ hybridization technique.</p><p><b>RESULTS</b>(1) The concentrations of MIP-1alpha in BALF and serum of group B(3) [(30.2 +/- 4.2) pg/ml, (30.8 +/- 4.6) pg/ml], group B(8) [(35.3 +/- 4.9) pg/ml, (34.9 +/- 5.1) pg/ml], group B(24) [(42.9 +/- 5.8) pg/ml, (41.7 +/- 6.3) pg/ml] and group B(36) [(37.8 +/- 4.7) pg/ml, (35.7 +/- 4.9) pg/ml] were significantly higher than those of group A(24) [(20.9 +/- 3.8) pg/ml, (22.4 +/- 4.3) pg/ml] (P < 0.01); the concentrations of MIP-1alpha in BALF and serum went up at 3 h, reached peak at 24 h, and had descended at 36 h. (2) Immunohistochemistry showed that the protein expressions of MIP-1alpha around the bronchus of group B(0) [(26.4 +/- 6.2)%] were significantly elevated as compared to those of group A(0) [(10.3 +/- 2.5)%] (P < 0.01), the epithelial cell was the chief expression cell. (3) In situ hybridization showed that the mRNA expressions of MIP-1alpha around the bronchus of group B(0) [(23.9 +/- 4.2)%] were significantly increased when compared to those of group A(0) [(8.7 +/- 1.8)%] (P < 0.01), the epithelial cell was the chief expression cell. (4) There was a significant correlation between the concentrations of MIP-1alpha and the numbers of EOS in BALF and between the concentrations of MIP-1alpha and the percentage of EOS numbers in the total cell numbers (EOS%) in BALF.</p><p><b>CONCLUSIONS</b>MIP-1alpha protein and MIP-1alpha mRNA were found strongly expressed in mouse asthma model, the epithelial cell was the chief expression cell; the kinetic characteristic of MIP-1alpha showed that its level increased at 3 h, reached peak at 24 h and declined at 36 h; MIP-1alpha and EOS gathering had a significant correlation.</p>

Eosinophils apoptosis, fas mRNA and bcl-2 mRNA expressions in asthma model of young rat and effects of achyranthes bidentata polysaccharides / 中华儿科杂志

<p><b>OBJECTIVE</b>Asthma is a chronic respiratory tract disorder characterized by airway hyperreaction (AHR), persistent airway inflammation, high serum IgE, overproduction of IL-4, IL-5 and IL-13 by allergen-specific Th2 cells. The morbidity and mortality of asthma have continued to increase despite the use of currently available therapeutic agents. The reputed effects of traditional Chinese medicines (TCMs) have led to increasing use of TCMs for treatment of asthma throughout the world. The aims of this study were to investigate in asthma model of young rat the mRNA expressions of apoptotic gene fas and bcl-2, eosinophils (EOS) apoptosis in airway, and effects of achyranthes bidentata polysaccharides (ABPS), a group of polysaccharides extracted from TCM Achyranthes bidentata blume, on treatment of asthma.</p><p><b>METHODS</b>Fifty Sprague-Dawley (SD) rats were divided into five groups, 10 rats per group. Asthma in rats was induced by intraperitioneal sensitization and challenge with nebulized ovalbumin (OVA). A pretreatment with ABPS [50 mg/(kg x d)] was done according to three different schedules: consecutively 3 days at sensitization (T1), at challenge (T2) or both of the two periods (T3). Sham-treated rats (A) and naive rats (C) served as controls. The animals were sacrificed 24 hours after the last challenge. The mRNA expression of bcl-2 and fas in eosinophils presenting in airway and the apoptosis of eosinophils in airway were assessed by using in situ hybridization with oligonucleotide probe and TUNEL methods, respectively.</p><p><b>RESULTS</b>(1) Twenty-four hours after the last antigen challenge, the mRNA expression of fas in eosinophils presenting in airway significantly decreased in group A [(43.4 +/- 10.0)%] compared with that in group C [(73.2 +/- 11.9)%] (P < 0.01). ABPS could increase the fas mRNA expression significantly in all the three groups [(59.0 +/- 8.1)%, (57.5 +/- 9.6)%, (76.2 +/- 2.7)%], compared with that in group A (P < 0.05, P < 0.05, P < 0.01, respectively). The expression of the bcl-2 mRNA in group C was (47.9 +/- 8.7)%, it was elevated to (67.4 +/- 7.3)% in group A (P < 0.01). The expression of the bcl-2 mRNA in ABPS treated T1 and T3 groups was significantly lowered [(57.7 +/- 12.7)%, (57.3 +/- 6.8)%, P < 0.05], but not in T2 group [(72.4 +/- 6.7)%]. (2) In group A, the EOS presenting in the airway increased significantly, but there were few apoptotic EOS; the percentage of apoptotic eosinophil was distinctly lower in group A than that in group C [(5.3 +/- 2.2)% vs. (15.9 +/- 2.4)%, P < 0.01]. Compared with that in group A, the eosinophil apoptosis ratio in those ABPS treated groups T1, T3 was evidently elevated [(8.7 +/- 2.9)%, (9.8 +/- 2.2)%, P < 0.05, P < 0.05], but ABPS treated at challenge (T2) could not change the eosinophil apoptosis ratio significantly (P > 0.05).</p><p><b>CONCLUSION</b>(1) In asthmatic rat, the expressions of the genes fas and bcl-2 mRNA in EOS were changed evidently and the ratio of EOS apoptotosis reduced greatly. (2) ABPS could enhance the apoptosis of EOS by upregulating the expression of the genes fas and bcl-2 mRNA.</p>