We have developed a general expression system that enhances the production
and secretion of proteins in mammalian cells. The protein of interest is
expressed as a fusion to a signal peptide and the Fc fragment of
immunoglobulin as the N-terminal fusion partner, which can direct the
cellular processes into expressing and secreting high levels of many
different types of proteins. These include secretory proteins, enzymes and
soluble domains of membrane proteins, as well as nuclear and regulatory
proteins. Typical expression levels of these proteins from stable cell
lines ranged from several to 100 microg/ml in conditioned media. The Fc
domain helps to solubilize hydrophobic proteins and provides a handle for
easy detection and purification of the fusion proteins; and it can be
cleaved off by treatment with protease if desired.
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OBJECTIVE: To determine whether blood lactate, base deficit, or oxygen-derived hemodynamic variables correlate with morbidity and mortality rates in a clinically-relevant LD50 model of penetrating trauma. DESIGN: Prospective, controlled study. SETTING: University research laboratory. SUBJECTS: Anesthetized, mechanically-ventilated mongrel pigs (30+/-2 kg, n = 29). INTERVENTIONS: A captive bolt gun delivered a penetrating injury to the thigh, followed immediately by a 40% to 60% hemorrhage. After 1 hr, shed blood and supplemental crystalloid were administered for resuscitation. MEASUREMENTS AND MAIN RESULTS: After penetrating injury, 50.7+/-0.3% hemorrhage (range 50% to 52.5%), and a 1-hr shock period, seven of 14 animals died, compared with six of six animals after 55% to 60% hemorrhage, and 0 of nine animals after < or =47.5% hemorrhage. Only two of 13 deaths occurred during fluid resuscitation. At the LD50 hemorrhage, peak lactate concentration and base deficit were 11.2+/-0.8 mM and 9.3+/-1.5 mmol/L, respectively, and minimum mixed venous oxygen saturation, systemic oxygen delivery, and systemic oxygen consumption were 33+/-5%, 380+/-83 mL/min/kg, and 177+/-35 mL/min/kg, respectively. For comparison, baseline preinjury values were 1.6+/-0.1 mM, -6.7+/-0.6 mmol/L, 71+/-3%, 2189+/-198 mL/min/kg, and 628+/-102 mL/min/kg, respectively. Of all the variables, only lactate was significantly related to blood loss before and after fluid resuscitation in the 16 survivors. However, r2 values were relatively low (.20 to .50), which indicates that only a small fraction of the hyperiactacidemia was directly related to tissue hypoperfusion. In the whole population of survivors and nonsurvivors, both lactate and base deficit (but none of the oxygen-derived variables) correlated with blood loss. CONCLUSIONS: Arterial lactate is a stronger index of blood loss after penetrating trauma than base deficit or oxygen-derived hemodynamic variables. The reliability of arterial lactate depends on several factors, such as the time after injury, the proportion of survivors and nonsurvivors in the study population, and on factors other than tissue hypoxia. 相似文献
OBJECTIVES: Invasive penile and urethral tumors are traditionally treated with aggressive excision that requires involved organ and adjacent organ sacrifice. An alternative approach seeks to completely excise the tumor with adequate margins while preserving form and function of the organ. We present 6 patients who underwent such organ-sparing surgery. METHODS: Six selected cases (4 penile and 2 urethral) are presented with operative photographs and pertinent data. RESULTS: Three distal tumors of the penis were treated with excision limited to the glans with histopathologic findings of verrucous carcinoma, melanoma, and angiosarcoma. One patient with squamous cell carcinoma of the distal shaft refused partial penectomy and underwent a local wedge resection. A patient with locally advanced bulbourethral transitional cell carcinoma (TCC) refused cystourethrectomy and underwent an anterior urethrectomy and perineal urethrostomy. A 48-year-old woman with an adenocarcinoma contained in a very distal urethral diverticulum underwent simple diverticulectomy and excision of distal urethra. Postoperative voiding and sexual function were well preserved. Follow-up was 12 to 48 months. The patient with angiosarcoma died of lung metastases at 48 months with no local disease, and the patient with bulbourethral TCC developed pelvic disease at 12 months with no local recurrence and died of metastases at 25 months. CONCLUSIONS: Organ-sparing surgery is appropriate in selected patients on the basis of stage and location, high risk of distant failure, and patient disposition. Close follow-up is necessary. Comanagement with reconstructive and oncologic specialists optimizes results and outcomes. 相似文献
A progressive fibrous myopathy may result from chronic intramuscular drug abuse. This complication may mimic other rheumatic disorders and early recognition may prevent disability. The patient described here presented with fixed flexion and extension contracture of hips and knees, respectively, after abusing meperidine and other agents for 3 years. Soft tissues of thighs and buttocks were "wood hard," EMG showed absence of action potentials in affected muscles, and biopsy revealed extensive replacement of muscle with dense, acellular fibrous tissue. Possible mechanisms are discussed. 相似文献
Studying the effects of various parameters on the behavior of velocity, temperature and thus the entropy generation rate in the microfluidic systems to reduce loss power is very important. Minimization of entropy generation in the flow system enables us for the parametric optimization of the MHD micropumps operation. In the present study, a transient, laminar and fully developed electrically conductive fluid flow in MHD micropumps has been investigated and the temperature distribution and effects of dimensionless influencing parameters on the entropy generation rate has been presented. Pumping operator in MHD micropumps are the Lorentz forces, which is produced as a result of the interaction between magnetic and electric fields. Governing equations have been solved numerically using finite-difference (ADI) method. The results of simulation have shown good agreement with analytical results by ei-genfunction expansion method. In addition, the results are compared with experimental data from literature which confirms the accuracy of the model. The obtained results showed that aspect ratio, Hartman, Prandtl, Eckert numbers and Joule heating parameter have significant influences on the flow and temperature distribution as well as entropy generation rate in MHD micropumps that controlling them can lead us for optimized operation of MHD micropumps.
The gls genes of Volvox are required for the asymmetric divisions that set apart cells of the germ and somatic lineages during embryogenesis. Here we used transposon tagging to clone glsA, and then showed that it is expressed maximally in asymmetrically dividing embryos, and that it encodes a 748-amino acid protein with two potential protein-binding domains. Site-directed mutagenesis of one of these, the J domain (by which Hsp40-class chaperones bind to and activate specific Hsp70 partners) abolishes the capacity of glsA to rescue mutants. Based on this and other considerations, including the fact that the GlsA protein is associated with the mitotic spindle, we discuss how it might function, in conjunction with an Hsp70-type partner, to shift the division plane in asymmetrically dividing cells. 相似文献
A number of studies have provided evidence that lipoprotein(a) [Lp(a)]
assembly is a two-step process in which initial non-covalent interactions
between apolipoprotein(a) [apo(a)] and apolipoproteinB-100 (apoB-100)
precede specific disulfide bond formation. We have designed a construct
encoding apo(a) kringle IV type 9 (KIV9) in which the unpaired cysteine at
position 67 in this kringle is replaced with a tyrosine. The single kringle
was expressed in bacteria and purified to homogeneity from cell
homogenates. The purified derivative (designated KIV9deltaCys) was assessed
for its ability to bind to purified human LDL. This interaction was
detected either by ELISA using immobilized LDL or by column chromatography
in which LDL binding to KIV9deltaCys immobilized on Ni2+-Sepharose was
determined. In both cases, the interaction of KIV9deltaCys and LDL was
observed. Further, we demonstrated that the binding interaction was
sensitive to the addition of amino acids including lysine, the lysine
analogue epsilon- aminocaproic acid, arginine, phenylalanine and proline,
with arginine and lysine having the greatest inhibitory effect. Binding of
KIV9deltaCys to an immobilized apoB peptide spanning residues 3732-3745 of
apoB was also demonstrated by ELISA. As was the case for LDL, this binding
interaction was sensitive to the addition of arginine and lysine. Computer
modeling of KIV9 demonstrated an excellent fit with residues 3732-3738
(PSCKLDF) of the apoB peptide. The modeling predicts the presence of
overlapping lysine and phenylalanine-binding pockets in KIV9 which explains
the inhibitory effects of lysine, arginine and phenylalanine which were
observed in the binding assays. In summary, this study represents the first
demonstration that KIV9 can interact directly with LDL through non-covalent
interactions which may contribute to the first step of Lp(a) formation.
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