富血小板血浆诱导脂肪干细胞成骨作用的实验研究

目的探讨在体外培养中富血小板血浆（PRP）对脂肪干细胞的增殖及诱导成骨的影响。方法从Wistar大鼠自体动脉血中提取PRP，配制成条件培养液，并作用于培养状态的脂肪干细胞，MTT法检测细胞的增殖情况，Kaplow法染色检测细胞碱性磷酸酶表达情况，碱性磷酸酶活性检测，茜素红染色鉴定钙结节形成情况。结果脂肪干细胞经诱导培养后，PRP诱导组较对照组增殖明显（P〈0．01），碱性磷酸酶活性较对照组增高明显（P〈0．01），PRP诱导组碱性磷酸酶染色阳性，茜素红染色可见钙结节形成。结论体外培养时，PRP可促进脂肪干细胞的增殖并能诱导成骨分化，从而为骨组织工程提供一种新的方法。     

A virological study of human kidney explant cultures from renal allograft recipients.

Human kidney tissue obtained at pretransplant nephrectomy from 13 adults and 1 child with end stage renal disease was cultured to provide a source for attempted detection of viruses. Cultures were observed throughout their life span for cytopathogenic viruses. Supernatants from 13 cultures were tested for hemagglutinating viruses (including measles, rubella, influenza, parainfluenza, and mumps) using various species of red blood cells. Kidney cells were cocultivated with Vero and human foreskin cells and indirect immunofluorescence was attempted in one instance. No viruses were detected. Nevertheless 9 of the 14 patients developed infections with herpes group viruses post-transplant. Either replicating viruses were not present in the kidneys of the patients prior to transplant or the techniques used were not adequate to detect them However, the establishment of organ cultures from patients with chronic renal disease is practical and may be a useful method for further virological investigation.     

瘢痕疙瘩不同分型皮损的血流及血管状况检测

目的：探讨瘢痕疙瘩（K）不同分型皮损的血流及血管状况。方法：将5例K患者皮损分为浸润K、增生K（红）、增生K（紫）、增生K（白）及老化K五型,同部位同时进行超声及皮肤镜检查,并与4例正常对照皮肤进行比较。结果：浸润K、增生K（红）、增生K（紫）及增生K（白）瘢痕组织内呈相对或绝对缺血或缺动脉血状态,浅表血管或细小、或通行异常、或通行不畅;老化K组织内血流状况不一,浅表血管形态分布接近正常皮肤。结论：K皮损分型不同,其血流状态及浅部血管表现亦不同。     

Physiologic effects of transfusing red blood cells with high or low affinity for oxygen to passively hyperventilated, anemic baboons: systemic and cerebral oxygen extraction.

Anemic, passively hyperventilated baboons were given preserved red blood cells either with increased or with slightly reduced affinity for oxygen to restore the red cell volume. In the high affinity group there was a 50% increase in cerebral blood flow immediately after the transfusion, but there was no significant change in the low affinity group. The cardiac output decreased slightly in the low affinity group, and increased slightly but insignificantly in the high affinity group. Two hours after transfusion the cerebral blood flow had returned to normal in the high affinity group. In both groups there was a decrease in arterial blood pH and an increase in Po2 in blood from the pulmonary artery and the jugular vein after transfusion. A 40% restoration of the 2,3 DPG level occurred within 4 hours of the transfusion of red cells with high affinity for oxygen, and this rapid increase was associated with increases in blood pH and inorganic phosphorus levels. Preserved red cells with high affinity for oxygen and low 2, 3 DPG levels significantly increased the cerebral circulation during the 2-hour posttransfusion period. These findings lend support to the recommendation that preserved red cells with normal or elevated 2,3 DPG levels be administered to patients in hemorrhagic or septic shock, and to patients subjected to extracorporeal circulation during cardiac surgery in order to lessen the demand for increased blood flow and to ensure adequate tissue oxygenation during the postoperative period.     

Cell salvage in obstetrics: an evaluation of the ability of cell salvage combined with leucocyte depletion filtration to remove amniotic fluid from operative blood loss at caesarean section

During 27 elective caesarean sections, operative blood loss was collected and processed using the Haemonetics Cell Saver 5 and filtered by Pall RC 100 leucocyte depletion filtration. The efficiency of removal of amniotic fluid, and the degree. of contamination with fetal red cells were assessed in the resulting 'cleaned' blood. Cell saver processing effectively removed alpha-fetoprotein from the red cells of 14 patients whose amniotic fluid was removed by separate suction and from nine of the 13 patients whose amniotic fluid was aspirated into the cell saver along with operative blood loss. Cell saver processing and leucocyte depletion filtration completely removed trophoblastic tissue and white cells, but fetal squames were still clearly present in 10, and possibly in 14 samples after processing and fully removed in only two specimens. Amorphous debris was present in all samples after processing. The maximum mass of fetal red cells contaminating any patient's total salvaged blood was 19 ml (range 2-19 ml). Had this been re-transfused into a rhesus-incompatible mother it would have required 2500 i.u. (500 microg) anti-D immunoglobulin to prevent rhesus-immunization of the mother. Contamination of processed caesarean section blood with fetal red cells and fetal squames is defined and its clinical implications discussed, with an overview of the development and current status of cell salvage. Autotransfusion by cell salvage with leucocyte depletion filtration should be considered in life-threatening obstetric haemorrhage and offered to Jehovah's Witnesses.     

A new automated system for urine analysis: a simple, cost-effective and reliable method for distinguishing between glomerular and nonglomerular sources of haematuria.

OBJECTIVE: To determine the ability, reliability and accuracy of a new automated system of urine analysis in differentiating glomerular from nonglomerular bleeding in the initial investigation of haematuria, and compare its efficacy with conventional phase-contrast microscopy (PCM). PATIENTS AND METHODS: One hundred and six urine samples from patients in whom the final diagnosis was available were analysed using electrical flow impedance to detect, enumerate and size red blood cells in a conductive fluid (the cellfacts analyser, Microbial Systems Ltd, Coventry, UK). All the samples were also tested using a dipstick method and PCM was carried out for comparison on 45 of the 106 urine specimens. The results of cellfacts analysis were correlated with the final diagnoses to assess sensitivity, specificity, positive predictive value (PPV), and negative predictive value (NPV) of this method; the costs were also analysed. RESULTS: Sixty-nine urine samples tested positive for blood on dipstick urine analysis and all these were confirmed to have red blood cells on cellfacts analysis. The remaining 37 samples were negative for blood on dipstick testing and cellfacts analysis, although seven patients had been referred with previously detected microscopic haematuria, none of whom were found to have any detectable pathology in the urinary tract on clinical examination and investigations. The remaining 30 patients were diagnosed to have urological or nephrological conditions with no haematuria. In the positive group, 20 (29%) patients were from the glomerular group, with a mean (range) red blood cell size of 4.25 (4-5.1) micrometer, and 49 (71%) from the nonglomerular group, with red blood cells of 5.47 (4.67-5.70) micrometer. These ranges overlapped at 4.67-5.1 micrometer at the decision threshold of 4.75 micrometer, the distribution of dysmorphic and eumorphic red blood cells for the glomerular group was 18 (90%) and two (10%), respectively, and for the nonglomerular group was 2 (4%) and 47 (96%), respectively. The sensitivity, specificity, PPV and NPV were 90%, 96%, 90% and 96%, respectively. Consumable and labour costs were very low. CONCLUSIONS: Cellfacts analysis is a simple, rapid, objective and cost-effective method for differentiating glomerular from nonglomerular urinary red blood cells, especially when few such cells are present.     

Intraoperative autotransfusion is associated with modest reduction of allogeneic transfusion in prosthetic hip surgery

Background: The efficacy of intraoperative salvage and washing of wound blood and the predictors of allogeneic red cell transfusions in prosthetic hip surgery are insufficiently known.
Methods: In 96 patients, undergoing primary or revision surgery, salvaged and washed red cells and, if necessary, allogeneic blood were used to keep haematocrit not lower than 33%. The bleeding of red cells during hospital stay was calculated from the red cell balance. The preoperative red cell reserve (millilitres of red cells in excess of a haematocrit of 33%) was estimated and the difference between this volume and the total bleeding of red cells was retrospectively used to classify patients with regard to the need for red cells. Stepwise regression analysis was used to define patient-related variables associated with allogeneic blood transfusion.
Results: Preoperative knowledge of the type of operation (primary, revision), the preoperative red cell reserve, and the body mass could predict roughly half of the need for banked blood (r²=0.45). Only one-third of the total bleeding of red cells was retransfused. For complete avoidance of allogeneic blood, autotransfusion was most effective in patients with a moderate need (0–4 u). However, 32% of such patients required allogeneic blood.
Conclusions: Autotransfusion has a limited efficacy to decrease the need for allogeneic blood, and other blood-saving methods should be added for this purpose. It is difficult to predict the need for allogeneic blood preoperatively.     

Enhanced regeneration of transplanted splenic tissue by increased work load to the splenic compartments

Autologous splenic tissue regenerates after subperitoneal transplantation in laboratory animals and in man. Qualitatively it resembles normal splenic tissue but the quantity usually only attains a small proportion of the normal spleen. In the prevention of overwhelming post-splenectomy sepsis a critical mass and a considerable blood flow to the regenerated spleen seem to be essential. By increasing the work load to splenic transplants in rats, significantly more splenic tissue was regenerated. This was achieved by: stimulating the white pulp by repetitive injections of xenogeneic red cells; stimulating the red pulp by damaging red cells with phenylhydrazine; a combination of 1 and 2; and stimulating the reticuloendothelial system by IP injections of methylcellulose. Stimulating the red pulp and the reticuloendothelial system were more effective than the injection of antigens. As the splenic mass is obviously regulated by the work load, we conclude that this effect should be used to attain the critical splenic mass and to increase the blood flow for effective clearance of bacteria from the blood.     

Prestorage leukoreduction prevents accumulation of matrix metalloproteinase 9 in stored blood

OBJECTIVES: Transfusion of aged stored blood is associated with many neutrophil proinflammatory effects. The mechanism of these effects remains to be elucidated. The purpose of this study was to determine whether matrix metalloproteinases accumulate in packed red blood cells during storage and are responsible for some of the neutrophil proinflammatory events, and to determine whether prestorage leukoreduction prevents accumulation of matrix metalloproteinases and attenuates proinflammatory effects of stored packed red blood cells. DESIGN: Laboratory study. PARTICIPANTS: Healthy human volunteers. INTERVENTIONS: Units of blood were drawn from healthy volunteers. Half of each unit was filtered for leukoreduction, removing 99.9% of leukocytes. At biweekly intervals, aliquots were removed from packed red blood cell units, and the plasma fraction was isolated for assays. Plasma was assayed for specific molecules or incubated with isolated neutrophils, with or without a matrix metalloproteinase 9 inhibitor. MAIN OUTCOME MEASURES: Concentrations of matrix metalloproteinases 2 and 9 and tissue inhibitor of metalloproteinase 1; matrix metalloproteinase 9 activity; and neutrophil apoptosis. RESULTS: Concentrations of matrix metalloproteinase 9 and tissue inhibitor of metalloproteinase 1 but not matrix metalloproteinase 2 increased over time. This accumulation was abolished by leukoreduction. Matrix metalloproteinase 9 accumulated in an active form. Both leukoreduction and matrix metalloproteinase 9 inhibition reversed stored packed red blood cell-induced, delayed neutrophil apoptosis. CONCLUSIONS: Storage of packed red blood cells for 14 days or more is associated with increases in the concentrations of matrix metalloproteinase 9 and tissue inhibitor of metalloproteinase 1, with the enzyme in excess of its inhibitor. Prestorage leukoreduction prevents this accumulation. Delayed neutrophil apoptosis related to packed red blood cell plasma appears to be due, in part, to matrix metalloproteinase 9. Leukoreduction can help prevent the effects of matrix metalloproteinase 9 on neutrophil apoptosis.     

Giant hemangioma of the tongue: combined use of perioperative blood conservation procedures

A giant hemangioma of the tongue was resected in a 16-year-old otherwise healthy young man (ASA I). Despite a total blood loss of 4,300 ml, corresponding to 105% of the patients intravascular blood volume, no allogeneic red blood cells had to be transfused intraoperatively. Besides minimization of intraoperative blood loss with preoperative alcohol injections into the tumor, ligation of large tumor-perfusing arteries, application of fibrin glue, skillful surgical technique, positioning of the surgical field above the level of the heart, controlled hypotension and maintenance of normothermia, acute normovolemic hemodilution (augmented by preoperative administration of recombinant human erythropoetin - rhEpo) and autotransfusion of lost blood were used for recovery of autologous blood. Under the protection of hyperoxia, a decrease of the hemoglobin (Hb) concentration to 4.2 g/dl was bridged by extreme normovolemic hemodilution. No signs of immanent or manifest tissue hypoxia were encountered. Retransfusion of autologous red blood cells was only started when surgical control of bleeding was achieved. Additionally a total of 4 units of fresh frozen plasma were infused for stabilization of plasma coagulation.After a 9-hour surgical duration, the patient was transferred to the intensive care unit, normotensive (with low-dose infusion of norepinephrin) and normothermic with a Hb concentration of 5.6 g/dl. In the face of an increasing lactacidosis 2 units of packed red blood cells were transfused on post surgical day 1.     

A comparison of transfusion requirements between living donation and cadaveric donation liver transplantation: relationship to model of end-stage liver disease score and baseline coagulation status

The use of living donation is an important option for patients in need of liver transplant. We retrospectively reviewed the preoperative Model for End-Stage Liver Disease (MELD) score, baseline coagulation laboratory results, and intraoperative transfusion of red blood cells and component therapy for 27 living donation transplants and 69 cadaveric donation transplants during a 3-yr period (2001-2004). Patients undergoing living donation transplantation had significantly lower MELD scores and preserved coagulation function compared with cadaveric donation transplantation recipients (P < 0.001). The living donation transplant patients also received significantly fewer transfusions of red blood cells and component therapy compared with the cadaveric donation transplant patients (P < 0.001). For the combined population of both cadaveric donation transplant and living donation transplant patients, there were significant associations between MELD score and preoperative coagulation tests (P < 0.001) and intraoperative transfusion of blood and component therapy. MELD score and preoperative fibrinogen concentration were identified as independent predictors of transfusion exposure. In conclusion, we detected significant differences in severity of disease at time of transplantation, degree of impairment of coagulation function, and need for transfusion of red blood cells and component therapy between patients undergoing living donation transplantation compared with patients undergoing cadaveric donation transplantation.     

The mechanisms of the changes in the erythrocyte oxygen-transport function of patients with chronic kidney failure undergoing treatment by systematic hemodialysis

Utilization by red blood cells of macroergic phosphates during cell incubation in polyionic medium was investigated in 15 healthy subjects and 20 glomerulonephritis patients on programmed hemodialysis for terminal chronic renal failure. By anemia severity the patients were divided into 2 groups. In less severe anemia there was more active utilization in glycolysis of 2,3-DPG and ATR in the presence of manifest oxidation of intraerythrocytic medium typical for Root effect predominance. This indicates that erythrocytic involvement into tissue oxygenation appears inadequate to high levels of macroergic phosphates. Hemodialysis in such patients promotes stabilization of red blood cell metabolism and Root effect suppression, thus contributing to maintenance of blood oxygen transport. In more severe anemia hemodialysis fails to establish biochemical grounds for improving erythrocytic gas transport function. The pattern of hemoglycolysis changes registered in cell incubation urge introduction of drugs stabilizing erythrocytic metabolism in the treatment of chronic renal failure.     

Red blood cell and tissue water content in experimental thermal injury

Oedema formation and changes in local blood flow are known phenomena in burns. The relationship between these two is not clearly described. The aim of this study was firstly to examine both the contents of red blood cells and tissue water in skin and subcutaneous fat after experimental burns of different depths in pigs, and secondly, to confirm our recent findings of the increased dielectric constant of skin and subcutaneous fat reflecting considerable oedema formation, especially in fat after thermal injury.

Methods

Superficial, partial and full thickness contact burns were created to pigs and followed for 24 h. Radioactive Cr-51 labelling of red cells was used to estimate the number of red cells in tissue, and the absolute amount of water was determined by lyophilisation.

Results

A decreased number of labelled red cells in skin and an increase in tissue water in subcutaneous fat were found regardless of burn depth. The highest water amount in fat was found in the partial thickness burns.

Conclusion

All burn depths resulted in a diminished number of labelled red blood cells in skin and a significant increase in the absolute water amount in subcutaneous fat at 24 h post injury. The findings in fat support our recent findings of highly elevated dielectric constants measured by the new in vivo method of dielectric measurements.     

The metabolic effects of fresh versus old stored blood in the priming of cardiopulmonary bypass solution for pediatric patients

OBJECTIVES: Pediatric cardiopulmonary bypass involves the creation of a large obligatory priming reservoir. Packed red blood cells are an essential part of the cardiopulmonary bypass priming solution in children. The storage media in packed red blood cells might cause significant acid-base, glucose, and electrolyte imbalances, which have been associated with severe complications. The purpose of the present study was to evaluate the metabolic effects of fresh (< or =5 days) versus old (>5 days) stored packed red blood cells added to the priming solutions of pediatric patients undergoing cardiac surgery. METHODS: Blood samples were drawn from cardiopulmonary bypass priming of 30 consecutive pediatric patients undergoing cardiac surgery. Patients were divided into 2 groups. Fresh (< or =5 days old) stored packed red blood cells were added to the priming solution in group 1, and old (>5 days old) stored packed red blood cells were added to the priming solution in group 2. In each group blood samples were drawn from the packed red blood cells on arrival to the operating room and from the priming solution immediately after packed red blood cells were added and after 20 minutes of prime circulation. Samples were also collected at the beginning of cardiopulmonary bypass and after 30 minutes. The last sample was collected on arrival to the pediatric intensive care unit. The levels of potassium, glucose, and lactate and the acid-base balance were analyzed in each sample. RESULTS: There was a linear increase in potassium levels in packed red blood cell samples with increasing packed red blood cell age, ranging from 5.4 to 18.4 mEq/L. Significant differences in the concentrations of potassium, glucose, and lactate and the acid-base balance were found when comparing old and fresh packed red blood cells in samples taken during the packed red blood cell and early prime time. Those differences resolved after 20 minutes of reconstitution of the priming solution. The age of the packed red blood cells had no effect on the samples taken during bypass and those taken in the pediatric intensive care unit. CONCLUSION: The significantly higher concentration of potassium and lactate and lower pH in old stored packed red blood cells has a minimal effect on the final constitution of priming solution before and during cardiopulmonary bypass in children undergoing corrective cardiac surgery.     

Autotransfusion in hepatectomy for hepatocellular carcinoma]

Experimental data of autotransfusion in hepatectomy for hepatocellular carcinoma and clinical data from forty-eight patients were analyzed. By a density gradient medium, cancer cells banded at a lower density (1.056 g/ml: mean) than red blood cells (1.105 g/ml: mean). The centrifuge autotransfusor separated the red blood cells from cancer cells and all sections of cell blocks of the blood harvested intraoperatively in 20 cases revealed no malignant cells. The average requirement of bank blood was significantly reduced in these patients than in patients without autotransfusion during one year before, 1573 ml to 478 ml (whole blood of concentrated red blood cells) (p less than 0.05). These findings indicate that autotransfusion in hepatectomy for hepatocellular carcinoma is safe and practical.     

The influence of extracorporeal circulation and hemoseparation on red cell deformability and membrane proteins in coronary artery disease

Extracorporeal circulation and hemoseparation may lead to coupled mechanical and chemical blood trauma and thus influence red cell deformability. Ten patients with coronary artery disease underwent coronary bypass. Patients' blood samples were drawn preoperatively, after extracorporeal circulation, and after hemoseparation. Ten healthy adults served as control subjects. Red blood cell deformability was determined by direct microscopic observation of red blood cells subjected to shear stresses of 1.2 to 13.3 Pa with a counter-rotating rheoscope. Red cell membrane proteins were separated by one-dimensional polyacrylamide gel electrophoresis in the presence of sodium dodecyl sulfate. At 1.2 Pa, preoperative red cell deformability was significantly greater in patients with coronary artery disease than in control subjects. Neither extracorporeal circulation nor hemoseparation changed red cell deformability significantly. Electrophoretic separation of membrane proteins failed to show any quantitative or qualitative differences between patients and control subjects. Moreover RBC membrane proteins of red blood cells in the patients were not altered as a result of extracorporeal circulation or hemoseparation. The preoperatively increased red cell deformability in the patients may be drug-induced. Our data suggest that the extracorporeal circulation and hemoseparation techniques used in this study do not lead to red blood cell damage.     

Evaluation of oxidative stress after repeated intravenous iron supplementation

Parenteral iron has been recommended for the treatment of iron deficiency in the majority of maintenance hemodialyzed (HD) patients. However, iron supplementation and consequent over saturation of transferrin and high iron levels, may aggravate oxidative stress already present in these patients. This study aimed to further clarify the role of repeated intravenous iron therapy as a supplementary cause of oxidative stress in HD patients. Markers of free radical activities (carbonyl reactive derivatives, CRD, thiol groups, SH, malondialdehyde, MDA) and antioxidant enzyme activities (superoxide dismutase, SOD and glutathione peroxidase, GPX) were determined in plasma and red blood cells (RBC) of 19 hemodialysis patients given a total iron dose of 625 mg (ferrogluconat, Ferrlecit, 62.5 mg). Blood samples were taken before the first and after the last dose of iron. Twenty apparently normal subjects served as healthy controls. Before iron treatment, HD patients exhibited increased concentrations of MDA and CRD in plasma and red blood cells, accompanied with impaired antioxidant capacity. All patients responded to iron therapy with a significant increase in their serum ferritin, serum iron, hemoglobin, and red blood cells levels. However, iron treatment resulted in enhanced oxidative stress in plasma of HD patients, since significant increase in plasma MDA and CRD concentrations, together with a decrease in nonprotein SH groups levels were detected. Supplementation with iron did not significantly influence plasma SOD and GPX activities, nor did any of the red blood cell parameters tested. Our data show that, despite improvement in hematological parameters, an increase in iron stores due to supplementation could also contribute to increased free radical production in HD patients.     

Further evidence supporting a cause and effect relationship between blood transfusion and earlier cancer recurrence.

Studies of associations between perioperative blood transfusions and later recurrence of solid tumors have yielded conflicting results. A previous analysis of transfused patients suggested that recurrence was associated with transfusion of whole blood as opposed to red blood cell concentrates. Additional analyses were performed on patients with cancers of the colon, rectum, cervix, and prostate to determine if patients receiving whole blood, red blood cells only, or no transfusions had differing outcomes. Patients receiving 1 unit or more of whole blood had uniformly poor outcomes compared with nontransfused patients (p less than 0.001). In contrast, patients receiving only red blood cells had progressively worse recurrence and death rates with increasing numbers of transfusion, suggesting the presence of a dose-effect relationship. Employing multivariate techniques, blood transfusion of less than or equal to 3 units that included any whole blood were independently and significantly associated with earlier recurrence (p = 0.003) and death due to cancer (p = 0.02). Transfusions of less than or equal to 3 units of blood comprised solely of red blood cell concentrates were associated with no greater risk of recurrence than that seen in patients receiving no transfusion (p = 0.50). These results provide a potential explanation for the disparate results reported in studies of blood transfusion and cancer outcome. The marked difference in outcome seen between patients receiving a few units of red blood cells and comparable patients receiving even one unit of whole blood are consistent with the hypothesis that transfusion of stored blood plasma causes earlier tumor recurrence in some instances. Strategies for reducing these risks might include avoidance of whole blood transfusions when only 1-3 units are required, more conservative transfusion practice, use of autologous blood transfusions, and perhaps, use of red blood cells washed free of plasma and white cell debris. Clinical trials to test these hypotheses are urgently needed.     

Splenosis mimicking tumor recurrence in renal cell carcinoma: detection on selective spleen scintigraphy.

A 2-year-old boy had been operated on for a giant renal cell carcinoma including splenectomy because of disrupture of the splenic capsule. During a follow-up examination, 3 nodules were detected by ultrasound in the splenorenal area. This gave reason to suspect tumor recurrence. Considering the possibility of splenosis, a selective spleen scan using denatured red blood cells was performed as a final diagnostic step. This method confirmed the nodules as representing splenic tissue. Splenosis should be included in the differential diagnosis of solid masses in the postsplenectomy patient.     

Red Blood Cell Susceptibility to Hydrogen Peroxide (H2O2) Lysis in Chronic Hemodialysis Patients

To assess susceptibility to oxidant induced membrane damage, the red blood cell H₂O₂ hemolysis test was performed in fifty-three chronic hemodialysis patients. Uremic red blood cells were not more susceptible to 2.5% H₂O₂ lysis pre or postdialysis than were control red blood cells. Oxidant induced red blood cell lysis is a function of the strength of the oxidant and not a testable intrinsic defect of the uremic red blood cell.