Objective To clarify the morphological features, types, regional distributions and cell densities of gastrointestinal (GI) endocrine cells in various parts of the digestive track (DT) of Eumeces elegans. Methods Using immunohistochemical techniques of streptavidin-peroxidase(S-P) method. Results The endocrine cells found in the DT were round or oval, spindle, shuttle, pyramid, flask and bacilliform in shape. Three types of endocrine cells, namely 5-hydroxytryptamine (5-HT) positive cells, somatostatin (SS) positive cells and gastrin (GAS) positive cells were found. No substance P(SP)-, glucagon (GLU)-, pancreatic polypeptide ( PP)- and insulin(INS) positive cells were detected. 5-HT positive cells distributed throughout the whole DT at various densities and they were most predominant cell types in the DT. SS positive cells were detected only in stomach. GAS positive cells showed a restricted distribution and were only demonstrated in the pylorus and duodenum. In the DT of E. elegans the region with the highest degree of cell type heterogeneity was pylorus and all types of endocrine cell along the DT showed peak density in the pylorus as well. Conclusion Some common features of the distribution of different types of GI endocrine cells are found between E. elegans and other reptiles. This common trait may reflect the similarity in digestive physiology of various vertebrates. On the other hand, some species-dependent unique distribution features of endocrine cells in DT were also detected in E. elegans.

Objective To observe the developmental changes of projection and termination of proprioceptive afferent fibers in the mouse spinal cord. Methods Parvalbumin (PV) immunohistochemistry was used to label the proprioceptive afferents. Single and dual immunofluorescence histochemistry were used to examine the growth pattern of proprioceptive afferents and their relationships with motoneurons in the spinal ventral horn (VH). The stained sections were observed under a confocal laserscanning microscope. Results PV-like immunoreactive (LI) proprioceptive fibers first appeared in the dorsal column on embryonic (E) day 14, then entered the gray matter on El5 and reached the intermediate gray matter and VH more obviously on E16. The number and intensity of PV-LI proprioceptive afferent fibers and punctata increased in the VH with age and reached a maximum during earlier postnatal (P) period (P0-P7). After P14, the number and intensity of proprioceptive afferents gradually decreased. The proprioceptive terminals seemed to form close relationship with motoneurons from E17. Conclusion The present study indicates that the somatotopic organization of proprioceptive afferents in the spinal cord is established during the late embryonic and early postnatal stages. These results provide evidence for understanding the development of the reflex movements.

Objective To investigate the spatiotemporal expression patterns and the relationship of α-sarcomeric actin(α-SCA) ,α-smooth muscle actin(α-SMA) and intermediate filament protein desmin with the maturation of the prenatal and the neonatal mouse hearts. Methods Serial sections of the embryo mouse and the neonatal mouse hearts were immunostained with antibodies against α-SCA, α-SMA and desmin. Results Ventricle and outflow tract of embryonic day(ED) 9 heart showed stronger expression of α-SCA and α-SMA, but desmin expression was lower. In the atrium, the expressions of α-SCA and α-SMA were restricted to the dorsal and ventral walls. In the sinus venosus, only a few weakly stained α-SCA positive cells were detected. No desmin expression was found in the atrium and sinus venosus. The expressions of α-SCA, α-SMA and desmin were increased to their highest level at ED 12. The higher expression of α-SCA remained to the postnatal stages. After ED 12, the expressions of α-SMA and desmin gradually decreased in different parts of the heart, but their expressions in the right ventricle persisted longer. After birth,desmin expression was mainly concentrated in the Z lines of I bands and intercalated disks. Conclusion The presence of spatiotemporal differences in the expression of α-SMA and desmin reveals regional differences in cardiomyocyte maturation in various parts of the embryonic mouse heart. The right ventricle shows a relatively slow pace of maturation. The α-SMA may contribute to a peristaltoid contraction pattern of the embryonic myocardium with a slow shortening speed, and a relatively higher level of desmin is required for the maturation of the sarcomere.

Objective To investigate the changes of neurotransmitters and the relationship between neurotransmitters and nerve regeneration during the optic regeneration in zebrafish. Methods Using the classical retinotectal regeneration model and electromicroscopy, we observed the ultrastructural changes of synapses of the superficial fiber and gray stratum(SFGS) in the tectum. Results The morphological changes of synapses can be divided into 4 stages: 1. Synaptic degeneration at the early stage after lesion. 2. Regenerating optic nerve fibers entered the SFGS laminar of the tectum, the densities of large granule vesicles (LGV) and small granule vesicles(SGV) were increased. 3. Lots of synapses were formed, the densities of small round clear vesicles (SCV) and small flat clear vesicles(FCV)were increased dramatically .4. Morphological recovery and refinement of the retinotopic innervation. Conclusion Neurotransmitters might play an important role during the regeneration of optic nerve, and they exhibited their effects in a chronological way.

Objective To report a new method of fluorescent labeling technique in microarray studies: universal primer U2 labeling( UPL). The efficiency was compared of the UPL with that of random primer, restriction display labeling method and the reverse transcription coupled random primer spiking labeling method(RT-PSL). Methods Influenza viral RNA was labeled with both UPL and the conventional random primer labeling method as well as two other more laborious labeling methods( RD-direct and RD-incorporate), and hybridized with influenza virus oligonucleotide microarrays. The signals extracted from the microarrays were analyzed using SPSS 10.0 software. Results The fluorescent intensity, signal-to-noise ration(SNR), true positive ratio(TPR) of probes and labeling reproducibility of UPL were demonstrated to be higher than those of the Random primer approaches.Conclusion These results established that UPL is a valid new labeling protocol, which may have wide applications in the research and development of the microarray technology.

Objective To study the condition and method of hydrodynamics-based transgene(HDT) in rat fatty liver. Methods Inject different dosages and concentrations of green fluorescent protein plasmid pEGFP-C1 at different speeds, then collect 4 rats' liver leaves at different time points after injection and prepare their frozen section, finally observe and quantify the GFP expression with fluo rescence microscope at 488 nm excitation wavelength. Results Plasmid pEGFP-C1 concentration 33mg/L, injection speed 2ml/s, injection volume 8.5%of rat body weight, injected plasmid. After 6 hours of injection, GFP-positive cells rate of pedicel leaf is about 18%, left leaf about 14%, middle leaf about 12.5%;R3ight leaf about 10% and tail leaf about 8%. GFP begin to gradually reduce since 24 hours, until 72 hours almost no GFP-positive cells were checked in all liver leaves. Conclusion Hydrodynamics-based transgene can be applied to rat fatty liver, the appropriate conditions of this method are 33mg/L plasmid concentration, 8.5% rat avoirdupois, 2ml/s injection speed, and the suitable time to observe the proportion of GFP-positive cells is 6-24 hours after gene injection.

Objective Measuring the displacement and deformation of the fifth screw under different tensile force before or after adultal internal fixation with compressing steel plate for femoral fractuer by digital speckle correlation method.And analysis of the displacement and deformation after reducing the number of screws.Aimed at providing a theoretical basis for clinical femoral internal fixation with compressing steel plate about the number of screws. Methods Taking the antisepsis femoral specimen of grown-ups, embedding and fixing with tooth powder after removing the soft tissue, producing a fixed model in the 1/2 femur.Fixing steel plate below the lateral femur subperiosteal with five screws through the fourth floor of cortical bone on both sides of the fractures, samples were divided into 10 states, group A-J were measured by orders, under the rally of 400 N,500N , measuring the displayment of the fifth screw with CSS-44100-e-universal test machine for loading rally experiment ,and automatic data processing. Results For analysis of variance, the mean displacement measurements of the fifth screw under rally of 400N,500N, B-J group and A group have significant differences(P<0.05). The mean deformation measurements of the fifth screw under rally of 400N,500N, group A and group F-J group have significant differences(P<0.05);group A and B-E group have no significant differences.Conclusion In a certain range, the fracture displacement and the numuber of the screws show negative correlation. The screws near the fracture line for femoral compression plate internal fixation of femoral fractuer is easily broken because of the too large deformation.

Objective Using shotgun mass spectrometry to detect proteins probably contained in bone marrow stromal cells(BMSCs) conditioned medium. MethodsMixed with BMSCs conditioned medium was divided into two parts which is(>5kD and <5kD) by means of ultrafiltration. The two parts were used to culture neural stem cells(NSCs) separately, and the proportions of neurons, astrocytes and oligodendrocytes in the offsprings of NSCs were calculated, then the effective part that could regulate the differentiation of NSCs was detected by Shotgun mass spectrometry. Results The BMSCs conditioned medium which is >5kD could promote the NSCs differentiate into more neurons and oligodendrocytes. The SDS-PAGE of this part showed that the most proteins were above 14kD, then the protein bands were enzymed. In total, 456 proteins were identified by Shotgun mass spectrometry after all the protein bands were enzymed, there were 154 similar proteins, 17 hypothetical proteins and 56 unknown proteins. And in the rest of 229 proteins, most of them were cytoskeletal proteins, secreted proteins, signal transduction proteins, enzymes, transporter and so on. Conclusion Many proteins secreted by BMSCs could regulate the differentiation of NSCs so as to prove the protein components probably existed in the BMSCs conditioned medium.

Objective To investigate the influence of pinealectomy(Px)and melatonin (MLT) supplementation on CD4~+CD25~+Treg cells development and Foxp3 expression in rat thymus. Methods One hundred and twenty clean-grade male SD rats were divided into five groups: normal control group, sham pinealectomized group, pinealectomized group, Px +7.5 mg/(kg·d) MLT group, Px +15 mg/(kg·d) MLT group, and the thymus were taken out at the 4th week and the 8th week;Flow cytometric analysis was used to analyse the number of double positive cells in the thymus and peripheral blood;Semiquantitative RT-PCR and immunohistochemistry were applied to analyse Foxp3 expression of rat thymus. Results There was no difference of the number of CD4~+CD25~+ Treg cells in rat thymus between Px and normal/sham group, and the number increased dependently on time and dose after MLT supplementation. In rat peripheral blood the double positive cells significantly increased in models of Px at the 4th week, and then backed to normal level after MLT supplementation. The results showed no significant changes in all groups at the 8th weeek;At the 4th weeks, Foxp3 expression increased in the thymus of Px rats compared to nomal/sham group, which returned to normal level after MLT supplementation. On the other hand, Foxp3 expression showed no significant difference in all groups at the 8th weeek.Conclusion Px made no difference between the development of CD4~+CD25~+Treg cells in rat thymus, but resulted in significant increase of thymic output and Foxp3 expression. All the effects disappeared at the 8th week. MLT supplementation could reverse the abnormity.

Objective To observe how exogenous estrogen influences the distribution and the expression of NOS positive neurons in the supraoptic nuclei of hypothalamus in the overiectomized female rats. Methods The 2-3-month-old female rats(n=40) were selected as the healthy and nulli-copulatory experimental animals. Rats were divided into following groups: normal control group(n=10), ovariectomized control group(n=10), and two experimental groups that have been injected with estrogen for post-operative 40 days(n=10) and for post-operative 70 days(n=10). Finally, all the animals were infused and the brains were removed. Immunohistochemical (SABC) method was adopted to count the number of NOS poitive neurons and observed the NOS poitive neuronal morphology under the light microscope. The image analysis system was used to test the average gray value of immunoreactivity in NOS positive neurons. Results In the ovariectomized control group, the density of NOS positive neurons in supraoptic nucleus was significantly increased and their shapes were bigger than that of the normal control group(P<0.05). The density and the form of the NOS positive neurons in supraoptic nucleons had no apparent difference between the estrogen for post-operative 40 days group and the ovariectomizeed control group(P>0.05).In the group after estrogen-injection 2 months compared with the normal control group, and the ovariectomized control group, both of the NOS positive neurons' density and the size become significantly decreased, and the staining of cells was lesser in the group injected with estrogen for post-operation 70 days. Conclusion The present results suggest that exogenous estrogen may influence the distribution and the expression of NOS positive neurons in supraoptic nucleus of hypothalamus of ovariectomized female rats.