Objective To construct a bait plasmid in bacterial two-hybrid system.Methods A cDNA fragment encoding for rat AMP-activated protein kinase ?2(AMPK?2) was amplified by PCR and inserted into bacterial expression vector pBT.After confirmation with restricted endonuclease digestion and sequence analysis,bacterial reporter strain XL-1 Blue MRF' was transformed with pBT-AMPK?2 plasmid and the expression of the recombinant bait fusion protein was detected.To test whether the bait fusion protein had the capability of self-activation,the XL-1 Blue MRF' cells were cotransformed with the pBT-AMPK?2 plasmid and empty pTRG vector,and screened on 3-amino-1,2,4-triazole(3-AT) Selective Screening Medium plates.Results Restriction digestion and sequence analysis revealed that the AMPK?2 code sequence was correctly inserted into pBT with a right reading frame.pBT-AMPK?2 expressed ?cI/AMPK?2 fusion protein.Colonies were obtained on no 3-AT Nonselective Screening Medium plates when XL-1 Blue MRF' cells were cotransformed with recombinant pBT-AMPK?2 and empty pTRG vector,while none grew on 3-AT plates,indicating that the recombinant plasmid pBT-AMPK?2 expressed AMPK?2/?cI fusion protein correctly,and was incapable of activation of the reporter cassette in the absence of an interaction partner.Conclusion The recombinant plasmid pBT-AMPK?2 could be used as "bait plasmid" to screen cDNA library.

With the advent of post-genomic era, identification of protein-protein interaction has become another hot spot of protein research and promoted the invention, development and complement of relative techniques. Major large-scale high-throughput methods, such as two-hybrid system, bacteriophage display, mass spectrometry, protein microchips and bioinformatics have provided a global view of protein-protein interaction, and are hopeful to play an important role in proteomics. Each method may have its specific strengths and weaknesses as well as differences in coverage, to some extent, the data based on these methods can complete each other. Here, recent progress of these large-scale high-throughput methods and their applications for studying protein-protein interaction are reviewed.

Objective To screen the proteins interacting with AMP-activated protein kinase ?2(AMPK?2)in rat brain cDNA library by bacterial two-hybrid system,and to investigate the biological role and the regulatory mechanism of AMPK?2 in brain.Methods The recombinant pBT-AMPK?2 was used as the bait to screen a rat fetal brain cDNA library by bacterial two-hybrid system.The plasmids of positive colonies were extracted and analyzed by DNA sequencing and BLAST search in GenBank.Results Seven proteins binding to AMPK?2 were screened out,including polyubiquitin,small heat shock protein 8,phosphofructokinase,cytochrome C oxidase subunit Ⅰ(COXⅠ),HLA-B-associated transcript 3(BAT3)isoform 1,protein tyrosine phosphatase receptor type D(Ptprd)and islet-brain 1(IB1).Conclusion Polyubiquitin,HSP8,phosphofructokinase,COXⅠ,BAT3 isoform 1,Ptprd and IB1 can interact with AMPK?2 in brain.

10.3969/j.issn.1000-3606.2013.06.003

AIM: To explore the effects of prostaglandi n I 2 (PGI 2) on mesenteric microcirculation and hemorheology during renal isch emia/reperfusion (IR) injury. METHODS: 36 rabbits were randomly distributed into the sham oper ated group (sham group), renal ischemia/reperfusion injury group (IR group) and PGI 2+IR group(PGI 2 group). IR group received clamping for 60 min follow ed by 120 min of reperfusion. A microcircular microscope image analysis system w as used to study the changes of mesenteric microcirculation and hemorheology at 60 min of ischemia and 120 min of reperfusion, respectively, while the blood sa mples were obtained for the measurement of hemorheological indexes. RESULTS: ① In IR group during the period of renal IR, the numb er of adhesive leukocytes and microthrombus, hemorrhage and hemorheological ind exes such as blood viscosity, plasma viscosity, blood reduction viscosity, he matocrit, erythrocyte aggregation index, erythrocyte sedimentation rate, eryt hrocyte sedimentation rate K and plasma fibrinogen were significantly higher, w hile microvascular diameters, blood flow velocity and erythrocyte deformation i ndex were significantly lower compared with sham group (P0.05). CONCLUSIONS: PGI 2 ameliorates the disturbance of mesenteric microcirculation and hemorheology caused by renal IR injury with the best effect at 10 ng?kg -1?min -1.

Objective To investigate the clinical features of patients with rhabdomyolysis and acute kidney injury (AKI) caused by intense exercise. Methods Data on patients with rhabdomyolysis and AKI due to intense military exercise from January 2002 to December 2017 in a single Chinese nephrology center were retrospectively reviewed. Parameters included clinical manifestations, markers of renal function and muscle damage, treatment and prognosis. Results Twenty-two male servicemen with AKI caused by rhabdomyolysis were included. They took part in the military running training before onset. 95.5％(21/22) took part in 5-kilometer race, of which cross-country was 86.4％(19 cases) and bare-handed was 9.1％(2 cases). Most cases occurred in summer, in which 72.7％(16 cases) took part in 5-kilometer cross-country race. The levels of serum creatinine (Scr), blood urea nitrogen (BUN), uric acid (UA) and creatine kinase (CK) significantly increased in patients, with 9.1％(2 cases) reaching AKI stage 1, 31.8％ (7 cases) reaching AKI stage 2, and 59.1％ (13 cases) reaching AKI stage 3, respectively. Serum CK levels were positively correlated with AKI stage (r=0.453, P<0.05), Scr (r=0.494, P<0.05) and BUN (r=0.545, P<0.01), while negatively correlated with UA (r=-0.487, P<0.05). Serum LDH levels were positively correlated only with age (r=0.533, P<0.05). Serum UA presented inverse correlations with BUN (r=- 0.513, P<0.05), K+ (r=- 0.642, P<0.01) and CK (r=- 0.487, P<0.05), and positive correlation with age (r=0.431, P<0.05). In particular, duration from onset of disease had a stronger positive association with BUN (r=0.907, P<0.01) and Scr (r=0.690, P<0.01). Of these patients with AKI, 21 cases(95.5％) reached complete recovery of kidney function and 1 case (4.5％) changed to chronic renal failure within 3 months after comprehensive treatments , including 8 cases(36.4％) who received appropriate continuous venovenous hemofiltration. Conclusions Intense exercise in summer is likely to cause rhabdomyolysis and AKI. Early diagnosis and comprehensive treatment including appropriate blood purification are crucial for a successful treatment. Our findings also emphasize the importance of age on muscle injury and the monitoring of electrolysts, markers of muscle damage and renal function for prevention of rhabdomyolysis and its related complications.

Objective:To investigate the current situation of medical social worker supervision in Shanghai, for reference to promote the high-quality development of medical social work.Methods:From June to July 2022, a questionnaire survey was conducted on the in-service medical social workers in all medical institutions with medical social work departments or posts in Shanghai. The questionnaire mainly included demographic information, current status of supervision implementation, and effectiveness of supervision. The data were analyzed descriptively with t test for comparison between groups and the Pearson test was for correlation analysis. Results:A total of 99 medical social workers were included in this study, 65 had received supervision, and medical institutions where 58 people located had established the supervision system. The average scores of actual and expected supervisory support received by medical social workers were 3.71 and 4.20 respectively, and the mean burnout level score was 32.91. The effect of establishing a supervision institutions on burnout was statistically significant ( P<0.05) and actual access to supervision support was negatively associated with burnout ( P<0.05). Conclusions:The overall situation of medical social work supervision in Shanghai was positive, but the demand for supervision was not fully satisfied; Supervision had a positive impact on reducing the burnout level. It is suggested that medical social work should further strengthen the cultivation of supervisory talents, promote the construction of supervisory systems, and improve the quality of supervision.