Pediatric acute respiratory distress syndrome (PARDS) is one of the most severe disease in pediatric critical care medicine with high mortality.Pediatric practitioners have recognized that ARDS in children is different from ARDS in adults.In the absence of identification of these differences,however,children have been characterized as having ARDS based on the adult definitions.Therefore,the managements for PARDS were conducted without specific considerations of children,and have limitations when applied to patients.With the purpose to highlight the gaps of ARDS between children and adults,the new insights into PARDS on the epidemiology,pathophysiology,diagnosis,treatment and prognosis in the recent years were summarized.

Objective To investigate the effect on differentiation of denervated skeletal muscle-derived stem cells (MDSCs) induced by TGF-β1 in vitro.Methods MDSCs were obtained from the rat denervated skeletal muscle by preplate technique,with TGF-β1 adding on medium.Cultured cells were divided into two groups.A,control group; B,10 ng/ml TGF-β1 group.Cell growth was observed with phase contrast microscope.lmmunocytochemistry,quantitative RT-PCR and Western blot was used to detect the expression of Sca-1,COL-Ⅰ,COL-Ⅲ,α-SMA and vimentin in denervated MDSCs.Results The synthesis of COL-Ⅰ,COL-Ⅲ,α-SMA and vimentin by denervated MDSCs was extremely low at protein level in vitro,while Sca-1 level was really high.Belong to the treatment with TGF-β1,COL-Ⅰ,COL-Ⅲ,oα-SMA and vimentin in the denervated MDSCs had strong expression,but Sca-1 in which had a weak expression.Under the stimulation of TGF-β1,COL-Ⅰ expression reached peak at the 2nd day (12.5591 ± 0.3389),which was about 3 times as control group.COL-Ⅲ reached highest value at the 5th day (0.8956 ± 0.0438),which was about 23 times as control group.α-SMA topped out to 18 times at the 5th day (1.1090 ± 0.0018).Vimentin expression rose by 8.5 times and peaked at the 5th day (0.1794 ± 0.0019).The expression of Sca-1 began to decline at the 2nd day,with a remarkable reduction at the 5th day (0.0636 ± 0.0015).Conclusion TGF-β1 could induce differentiation of the denervated MDSCs to myofibroblasts in vitro,and promote the synthesis and excretion of extracellular matrix.

BACKGROUND:Nowadays increasing experimental findings have proved that the low-frequency pulsed electromagnetic fields (LPEMF) can induce osteogenic differentiation of a variety of stem cells in the two-dimensional scaffold. However, little is reported on the LPEMF effect on the proliferation and osteogenic differentiation of stem cells in the three-dimensional scaffold.OBJECTIVE:To investigate the effect of LPEMF on the proliferation and osteogenic differentiation of human adipose-derived stem cells (hASCs) in the 3D Insert-PCL scaffold.METHODS:Passage 3 hASCs were directly cultured in the 3D Insert-PCL scaffolds folowed by LPEMF (50 Hz, 1 mT) exposure, 2 hours per day, for continuous 14 days (experimental group) or no intervention (control group). After 7 days of culture, Live/Dead staining was used to observe cell survival. After 1, 3, 5, 7 days of culture, MTT assay was used to detect cell proliferation. After 7 and 14 days of culture, the osteogenic differentiation of hASCs was assessed through the alkaline phosphatase (ALP) staining and qRT-PCR.RESULTS AND CONCLUSION: Live/dead cell staining proved that the hASCs had a good growth in the 3D Insert-PCL scaffolds as well as a high survival rate. The absorbance values of hASCs in the two groups were increased gradualy with time, and the absorbance value in the experimental group was significantly higher than that in the control group at 1 and 3 days after culture (P < 0.05). The ALP activity in the experimental group was stronger than that in the control group at 7 and 14 days after culture. qRT-PCR findings showed that at 7 days after culture, the mRNA levels of ALP and type Ⅰ collagen were significantly higher in the experimental group than the control group (P < 0.01), while at 14 days after culture, the mRNA levels of osteopontin, Runt-related transcription factor, ALP and type Ⅰ collagen were significantly higher in the experimental group than the control group (P < 0.05). To conclude, the LPEMF exposure can promote the proliferation and osteogenic differentiation of hASCs cultured on the the 3D Insert-PCL scaffold.

Objective To construct eukaryotic expression plasmid pIRESneo3-pigment epithelium-derived factor (PEDF) and detect its transient expression in SP2/0 cells. Methods Specific primers were designed based on the mature peptide sequence of human PEDF cDNA in the GenBank. Human PEDF gene was cloned into the eukaryotic expression vector pIRESneo3. The PEDF DNA was transfected into SP2/0 with LipofectamineTM 2000. The recombinant human PEDF protein expressed in SP2/0 cell culture supernatant was identified by Western blot and enzyme-linked immunosorbent assay. The biological activity of the recombinant human PEDF was measured by 3-(4,5-dimethylthiazol-z-y1)-2,5-diphenytetrazolium bromide(MTT) method. Results PCR amplification, restriction enzyme digestion and DNA sequencing confirmed that the mature peptide sequence of human PEDF cDNA was successfully cloned into the eukaryotic expression vector pIRESneo3. And the plasmid was transfected into SP2/0 cells, which could secret PEDF. Western blot analysis showed that there was only one obvious band at the position of relative molecular weight of 50 000, and it is equivalent to the expected value. Enzyme-linked immunosorbent assay suggested that the content of PEDF began to rise after transfection, and peaked at 36 h [(0.92±0.04) μg/ml]. The proliferation of human umbilical vein endothelial cell line was significantly inhibited by supernatant after transfection of 36 h (P<0.05). Conclusions The eukaryotic expression plasmid pIRESneo3-PEDF had been successfully constructed and active human PEDF was transiently secreted, which made a foundation for further study of stable expression and purification of PEDF. This protein could be a potential medication for preventing and managing retinopathy of prematurity.

Objective To evaluate the role of haeme oxygenase-1 (HO-1) in remote limb ischemic preconditioning (RLIP)-induced attenuation of lung ischemia-reperfusion (I/R) injury in rabbits.Methods Twenty-four Japanese White Rabbits,aged 4-5 months,weighing 2.0-2.5 kg,were randomly divided into 4 groups (n =6 each) using a random number table:sham operation group (S group),I/R group,RLIP group and zinc protoporphyrin (ZnPP,an inhibitor of HO-1) plus RLIP group (ZnPP + RLIP group).Lung I/R was produced by 60 min occlusion of the left lung hilum followed by 180 min of reperfusion in I/R,RLIP and ZnPP + RLIP groups.RLIP and ZnPP + RLIP groups received 3 cycles of 10 min ischemia followed by 10 min reperfusion in the bilateral hind limbs immediately before occlusion of the left lung hilum.In ZnPP + RLIP group,ZnPP 10 μmol/kg was injected intravenously 10 min prior to hind limb ischemia and the rest of the procedures were similar to those previously described in RLIP group.At the end of reperfusion,arterial blood samples were collected for blood gas analysis.The animals were then sacrificed and pulmonary specimens were obtained for microscopic examination of the pathological changes which were scored (lung injury score,LIS) and for determination of wet/dry lung weight ratio (W/D ratio),myleoperoxidase (MPO) activity,malondialdehyde (MDA) content and expression and activity of HO-1 in the lung tissues.Results Compared with group S,PaO2 was significantly decreased,and LIS,W/D ratio,MPO activity,MDA content,and HO-1 expression and activity were increased in I/R group (P ＜ 0.01).Compared with I/R group,PaO2 and HO-1 expression and activity were significantly increased,and LIS,W/D ratio,MPO activity and MDA content were decreased in RLIP group (P ＜ 0.01).Compared with RLIP group,PaO2 and HO-1 expression and activity were significantly decreased,and LIS,W/D ratio,MPO activity and MDA content were increased in ZnPP + RLIP group (P ＜ 0.01).Conclusion RLIP up-regulates HO-1 expression and enhances HO-1 activity,thus reducing lung I/R injury in rabbits.

In order to investigate the biological function of transforming growth factor-β1 (TGF-β1) during fibrosis in denervated skeletal muscle, we recruited sciatic nerve injury model of SD rats in which denervated gastrocnemius was isolated for analysis. At different time points after operation, denervated muscle was examined by several methods. Masson trichrome staining showed morphological changes of denervated skeletal muscle. Quantitative RT-PCR detected the rapid increase of TGF-β1 expression at mRNA level after nerve injury. It was found that a peak of TGF-β1 mRNA expression appeared one week post-operation. The expression of collagen I (COL I) mRNA was up-regulated in the nerve injury model as well, and reached highest level two weeks post-injury. Immunoblot revealed similar expression pattern of TGF-β1 and COL I in denervated muscles at protein level. In addition, we found that the area of the gastrocnemius muscle fiber was decreased gradually along with increased interstitital fibrosis. Interestingly, this pathological change could be prevented, at least partly, by local injection of TGF-β1 antibodies, which could be contributed to the reduced production of COL I by inhibiting function of TGF-β1. Taken together, in this study, we demonstrated that the expression of TGF-β1 was increased significantly in denervated skeletal muscle, which might play a crucial role during muscle fibrosis after nerve transection.

ObjectiveTo investigate expression of TGF-β1,CTGF and collagen deposition in skeletal muscle during chronic entrapment of peripheral nerve. MethodsFifty rats were separated into two groups,control group and experimental group. At different time points after operation, the right gastrocnemius of 5rats from each group were collected for further analysis such as HE, Masson stain, immunohistochemical staining,RT-PCR and Western-blot. Results It was observed that axon degeneration occurred during chronic nerve entrapment,and which was in line with reports from other groups.Moreover,it had been demonstrated that after nerve entrapment,skeletal muscles may form fibrosis and degeneration consequently.Within this pathological procedure,expression of TGF-β1. CTGF and deposition of collagenⅠ changed rapidly when compared with control group.ConclusionOverall,these results indicated that these factors may be important during skeletal muscle degeneration after chronic nerve entrapment.

To study the characteristics of acute rejection after limb allotransplantation, 29 male Sprague-Dawley rats were randomly divided into 2 groups, with 15 rats in control group and 14 rats in experimental group. Each rat in control group underwent limb replantation. Each rat in experimental group received limb transplantation from Wistar rat. No immunosuppressive drugs were used after operation. The circulation of the transplanted limb, time and signs of rejection, histopathological changes in the tissues of the limb graft when rejected and survival time of limb grafts were evaluated. In the control group, no signs of rejection were observed, the circulation of each replanted limb was normal, it could survive for a longer time. The experimental group showed clinical signs of rejection (sub dermal edema and erythema) after a mean time of 3.36+/-1.15 days, and the mean survival time of the allografts was only 7+/-0.78 days. Histopathological examination showed most violent rejection reaction in skin. It is concluded that with Wistar-to-SD limb transplantation without use of immunosuppression, rejection of the grafts would occur after a mean time of 3.36+/-1.15 days; the earliest signs of rejection were edema and erythema of the skin, skin being the most representative component of limb graft rejection.
Acute Disease ; Extremities/*transplantation ; *Graft Rejection ; Graft Survival ; Random Allocation ; Rats, Sprague-Dawley ; Rats, Wistar ; Skin/immunology ; Transplantation, Homologous

Protective effect of interleukin-1beta (IL-1beta) on motor neurons was studied after peripheral nerve injury. Twenty Wistar rats were divided into 2 groups randomly. The right sciatic nerve of each rat was resected. After silicon tubulization of sciatic nerve in rat, 15 microl 1 ng/ml IL-1beta and PBS solution were injected into the silicon capsule respectively. Enzyme histochemistry was performed to show acetyle cholesterase (AchE) and nitric oxide staining (NOS) activity of spinal alpha motor neurons in spinal segments 2 weeks later. Neurons were counted and the diameter and cross sectional (c/s) area of neurons were analyzed by using computer image analysis system. The results showed that as compared with the normal side, both enzyme activities significantly changed in motor neurons in PBS group. The diameter and c/s area of both neurons changed significantly too (P < 0.01). These results suggest that exogenous IL-1beta protects alpha-motor neurons from degeneration and necrosis after peripheral nerve injury.
Interleukin-1/*pharmacology ; Motor Neurons/*pathology ; Neuroprotective Agents/*pharmacology ; Random Allocation ; Rats, Wistar ; Sciatic Nerve/*injuries ; Spinal Cord/pathology

BACKGROUND: In repair of nerve defect with allogenic nerve graft, to reduce immune rejection is one of the key problems. At present, the main approach is to reduce antigenicity of grafted nerve segment and apply generally immune inhibitor.OBJECTIVE: To observe the effects of freeze/thaw treatment and local application of transforming growth factor beta 1 (TGF-β1) plasmid on frozen nerve allograft.DESIGN: Randomized controlled animal experiment was designed.SETTING: Department of Hand Surgery, Union Hospital, Tongji Medical College Affiliated to Huazhong University of Science and Technology.MATERIALS: The experiment was performed in Tongji Medical College of Huazhong University of Science and Technology from January 2003 to December 2004, in which 40 Wistar healthy and adult rats were employed,from different delivery and were randomized into experimental group and control, 20 rats in each one.METHODS: Transforming growth factor-β1 (TGF-β1) plasmid and frozen allogenic sciatic nerve were prepared. In experimental group and control,sciatic nerve was cut off 2.0 cm in length, in the foramen 0.5 cm beneath piriformis. The nerve defect was repaired with pre-frozen allogenic nerve 2.0 cm in length. In experimental group, TGF-β1 plasmid was injected in local muscle and two broken ends of nerve. In the control group, physiological saline of equal volume was injected. In the 6th and 12th weeks, the samples were collected from 10 rats in each group for sectioning, staining,axonal counting and statistical analysis.RESULTS: No any animal was died in experiment and all of animals entered result analysis. In the 6th weeks, in the control group, mild edema appeared among axons on the grafted segment of nerve and in the experimental group, there was no edema among axons and the regenerated nerve numbers were close to the normal. In the 12th week, in the experimental group, the entire grafted nerve segment was basically filled up by the regenerated axons;myelinated nerve fiber was arranged in order and both axons and myelins were developed well. The regenerated axonal count in experimental group was more significantly than the control, indicating extremely significant difference [(98.6±4.8), (75.8±5.1) counts/μm2, t=2.962, P ＜ 0.01].CONCLUSION: Freeze/thaw treatment can decrease antigenicity of allogenic nerve, which provides the possibility of repair of nerve defect. Local application of TGF-β1 plasmid can provide immune inhibition locally and reduce immune rejection in the host.