Long-term protection of hematopoiesis against the cytotoxic effects of multiple doses of nitrosourea by retrovirus-mediated expression of human O6-alkylguanine-DNA-alkyltransferase

A human O6-alkylguanine-DNA-alkyltransferase (ATase) cDNA-containing retrovirus was used to infect murine long-term primary bone marrow cultures. High levels of ATase expression were obtained, and colony- forming cells of the granulocyte-macrophage lineage from the cultures transduced with the human ATase retrovirus were three times more resistant to the alkylating agent, N-methyl-N-nitrosourea (MNU), than control cultures. Furthermore, expression of the human ATase protected long-term hematopoiesis, measured as the output of progenitor cells to the nonadherent fraction of the culture, against the cytotoxic effects of repeated exposures to MNU. These results clearly show that a human ATase cDNA-containing retrovirus can be used to infect long-term primary bone marrow cultures and that this attenuates their sensitivity to nitrosoureas.     

Enhancement of chemotactic factor-stimulated neutrophil oxidative metabolism by leukotriene B4

Leukotriene B4 (LTB4) is a potent primary stimulator of neutrophil chemotaxis, aggregation, and degranulation and induces superoxide production at higher concentrations. In order to determine whether LTB4 modulates neutrophil responses to oxidative stimuli, human neutrophils (PMNs) were incubated with LTB4 prior to stimulation with f-Met-Leu-Phe (fMLP, 10(-7) mol/L), opsonized zymosan (OZ, 250 micrograms/mL), or phorbol myristate acetate (PMA, 32 nmol/L). Superoxide (O2-) production by stimulated PMNs was assessed by the superoxide dismutase-inhibitable reduction of cytochrome c. LTB4 alone did not stimulate O2- production in concentrations below 10(-7) mol/L and had no effect on the O2- assay. In the concentration range of 10(-12) to 10(-8) mol/L, LTB4 did not alter O2- release induced by OZ or PMA. In contrast, LTB4-treated cells demonstrated enhanced O2- production following exposure to fMLP, and in the presence of 10 nmol/LLTB4, generated 180% +/- 41% of O-2 quantities produced by control cells (n = 23). Enhancement was LTB4 dose-dependent, was maximal in the range of 1 to 10 nmol/L LTB4, was not reversed by removal of the lipid from the medium prior to fMLP stimulation, and was not dependent on the presence of Ca++ or Mg++ in the suspending medium. Chemiluminescence of fMLP-stimulated neutrophils was increased to 323% of controls in neutrophils preincubated with 10 nmol/L LTB4. Unlike augmentation of oxidative responses to fMLP seen with other degranulating stimuli, enhancement by LTB4 was not correlated with an increase in 3H-fMLP receptor binding. These results indicate that, in addition to its primary effects on neutrophil function, LTB4 modulates PMN oxidative responses to the chemotactic peptide and, thus, may amplify the release of oxygen metabolites at inflammatory foci.     

von Willebrand disease "Vicenza" with larger-than-normal (supranormal) von Willebrand factor multimers

When normal volunteers or patients with type I von Willebrand disease (VWD) are given desmopressin (DDAVP), a set of larger-than-normal (supranormal) von Willebrand factor (VWF) multimers, similar to those present in VWF-containing cells such as platelets megakaryocytes and endothelial cells, appear transiently in postinfusion plasma. In two kindreds with mild lifelong bleeding symptoms transmitted as an autosomal dominant trait, all ten symptomatic members (but none of the five asymptomatic members) had a supranormal multimeric structure for plasma VWF, apparently identical to that seen for postdesmopressin normal plasma. Plasma factor VIII coagulant activity (VIII:C), VWF antigen (VWF:Ag), ristocetin-induced platelet agglutination, and ristocetin cofactor (RiCof) activity were low. Platelet VWF:Ag and RiCof levels (tested for three patients only) were normal. Bleeding times were normal or slightly prolonged. The patients' platelet multimeric structure was the same as that for normal platelets. After desmopressin infusion the plasma VWF multimeric structure remained supranormal as for preinfusion plasma, with VIII:C VWF:Ag and RiCof increasing markedly over baseline values and disappearing at a normal rate. Examination of the VWF subunit composition from three of these patients indicated that proteolytic processing of their VWF did not differ from normal. This study describes the first variant of VWD with a supranormal multimeric structure.     

Return to sporting activity after Birmingham hip resurfacing arthroplasty: Mid term results

Background:

Hip resurfacing arthroplasty (HRA) is primarily indicated for young, active patients with disabling coxarthrosis who wish to remain active and return to sports after surgery. Relatively few prospective studies have assessed return to sporting activity and impact of gender and age on this.

Materials and Methods:

Seventy-nine consecutive patients treated with HRA were included. Patients were reviewed clinically and radiologically. Function was assessed using the modified University of California Los Angeles (UCLA) activity score. The Oxford, Harris and WOMAC hip scores were calculated.

Results:

Average age at the time of surgery was 54.9 years (range 34.5–73.6 years). Average preoperative and postoperative UCLA scores were 4 and 7.6 respectively. Patients were involved in 2 (0–4) sporting activities preoperatively and 2 (0–5) postoperatively. Preoperative and postoperative Oxford Hip Scores, Harris Hip Score and WOMAC scores were 40, 46 and 51 and 16, 94 and 3 respectively (P < 0.0001). Patients returned to sports at an average of 3 months postoperatively.

Conclusion:

Patients were able to return to sports by 3 months and perform the same number of activities at preoperative intensity. Activity levels are maintained up to the medium term with few complications.     

Systematic review of enhanced recovery after gastro-oesophageal cancer surgery

Introduction

Fast track methodology or enhanced recovery schemes have gained increasing popularity in perioperative care. While evidence is strong for colorectal surgery, its importance in gastric and oesophageal surgery has yet to be established. This article reviews the evidence of enhanced recovery schemes on outcome for this type of surgery.

Methods

A systematic literature search was conducted up to March 2014. Studies were retrieved and analysed using predetermined criteria.

Results

From 34 articles reviewed, 18 eligible studies were identified: 7 on gastric and 11 on oesophageal resection. Three randomised controlled trials, five case-controlled studies and ten case series were identified. The reported protocols included changes to each stage of the patient journey from pre to postoperative care. The specific focus following oesophageal resections was on early mobilisation, a reduction in intensive care unit stay, early drain removal and early (or no) contrast swallow studies. Following gastric resections, the emphasis was on reducing epidural anaesthesia along with re-establishing oral intake in the first three postoperative days and early removal of nasogastric tubes.In the papers reviewed, mortality rates following fast track surgery were 0.8% (9/1,075) for oesophageal resection and 0% (0/329) for gastric resection. The reported morbidity rate was 16.5% (54/329) following gastric resection and 38.6% (396/1,075) following oesophageal resection. Length of stay was reduced in both groups compared with conventional recovery groups in comparative studies.

Conclusions

The evidence for enhanced recovery schemes following gastric and oesophageal resection is weak, with only three (low volume) published randomised controlled trials. However, the enhanced recovery approach appears safe and may be associated with a reduction in length of stay.     

A new variant of type II von Willebrand disease with aberrant multimeric structure of plasma but not platelet von Willebrand factor (type IIF)

A patient with a lifelong bleeding disorder was diagnosed as having Type II von Willebrand disease. The larger multimers of von Willebrand factor were absent from her plasma but present in platelets. A high- resolution electrophoretic technique was used to study the complex structure of individual von Willebrand factor multimers. In normal plasma, each multimer could be resolved into five bands: a more intense central one and four less intense, two moving faster and two slower than the central band. In normal platelets, each multimer could also be resolved into five bands. The central one had a mobility similar to that in plasma, whereas the four satellite bands had a mobility that differed from that of the corresponding plasma bands. In the patient, platelet von Willebrand factor antigen content and ristocetin cofactor activity were normal, and von Willebrand factor showed the same structure of individual multimers as seen in normal platelets. On the other hand, plasma von Willebrand factor antigen and ristocetin cofactor activity were decreased, and the structure of individual von Willebrand factor multimers was different from that of normal plasma and similar to that seen in normal and patient's platelets. After infusion of 1-deamino-8-D-arginine vasopressin, the largest von Willebrand factor multimers, as well as new satellite bands with a mobility similar to those in normal plasma, appeared in the patient plasma, and the levels of von Willebrand factor antigen and ristocetin cofactor activity became normal. Yet no relevant change in the prolonged bleeding time was observed. This new variant of von Willebrand disease, therefore, is characterized by the presence of a dysfunctional von Willebrand factor molecule that exhibits unique structural abnormalities in plasma but appears to be normal in platelets. The designation of Type IIF is proposed for this type of von Willebrand disease in accordance with the terminology that has been previously used.     

Hemostatic plug formation in normal and von Willebrand pigs: the effect of the administration of cryoprecipitate and a monoclonal antibody to Willebrand factor

Hemostatic plug (HP) formation was investigated in the ear bleeding time incision in normal and von Willebrand pigs. HP volume was calculated by integrating the areas of serial sections. In normal pigs (n = 11), platelets immediately formed a layer on the surface of the cut channel. Platelet aggregates formed at the ends of transected vessels and gradually enlarged. Finally, all transected vessels were occluded by HP and bleeding stopped. In contrast, large HPs were formed in the incision in von Willebrand's disease (vWD) pigs (n = 4); these HPs did not cover the ends of the transected vessels, which continued to bleed, allowing the formation of large hemostatically ineffective platelet aggregates in the incision. Canals traversed these HPs, and bleeding from the open vessels may have continued through them. After infusion of cryoprecipitate into a vWD pig, the bleeding time shortened, and the morphological findings of the HPs were similar to those of normal pigs. In normal pigs (n = 3) infused with an anti- Willebrand factor monoclonal antibody, which prolonged the bleeding time, a large HP formed in the incision, similar to that observed in the vWD pig. The volume of the normal and vWD HPs increased with time. These in vivo findings suggest that Willebrand factor is involved in the localization of the HP to the damaged vessel and may also play a role in platelet-platelet interaction. A computerized morphometric technique was used for measuring the volume of the hemostatic plugs and the distance of sequential points on the perimeter of the HP from the center of selected bleeding vessels.     

Effect of n-3 and n-6 fatty acids on proliferation and differentiation of promyelocytic leukemic HL-60 cells

Promyelocytic leukemic HL-60 cells were incubated with different fatty acids. Arachidonic acid (AA; 20:4, n-6) and eicosapentaenoic acid (EPA; 20:5, n-3) were the most potent inhibitors of proliferation in a dose- dependent way. Retinoic acid (RA) was used as a positive control. Inhibitors of cyclooxygenase and lipoxygenase or addition of antioxidants did not influence the effect of EPA or AA on cell proliferation. Increased capacity to generate superoxide anions after phorbol ester treatment and a reduced serglycin messenger RNA level in cells treated with AA or EPA indicated that these fatty acids induced differentiation in HL-60 cells similar to that induced by RA. However, down-regulation of the c-myc mRNA level, also typical for differentiation with RA in HL-60 cells, was not observed in cells incubated with AA or EPA. Flow cytometric analyses showed that in cultures incubated with AA or EPA, the proportion of cells in the G1 phase of the cell cycle increased. Similar effects were observed with RA. By flow cytometry and light scatter analyses it could be shown that AA made 8% of the cells apoptotic and 7% necrotic. The corresponding numbers were 21% and 10% for RA-treated cells, and 19% and 32% for EPA- treated cells. The present study shows that AA and EPA reduce the proliferation rate of HL-60 cells. This is mediated by mechanisms independent of eicosanoids or lipid peroxidation products and is due to effects both on apoptosis/necrosis and cell differentiation.     

Immunologic status of hemophilia patients treated with cryoprecipitate or lyophilized concentrate

We evaluated 37 patients with moderate or severe hemophilia A and six patients with severe factor IX deficiency for clinical or laboratory evidence of immune abnormalities. Patients were assigned to one of four groups according to the type of clotting factor replacement. Twenty patients had received only cryoprecipitate during the two years preceding the evaluation (group I); 11 additional patients were treated predominantly with cryoprecipitate but had also received up to nine bottles of factor VIII concentrate (group II); six patients received factor VIII concentrate (group III); six patients received factor IX concentrate (group IV). There was no clinical or laboratory evidence of immunodeficiency among the 43 patients. The mean absolute number of Th cells was normal in all patient groups, but the mean absolute number of Ts cells was increased compared with controls, both in patients treated with cryoprecipitate and in patients treated with factor VIII or factor IX concentrate. There was no correlation between the Th/Ts ratio and patient age, alanine aminotransferase level, hepatitis serology, in vitro lymphocyte function, or amount of clotting factor administered. Our observations demonstrate that the volunteer or commercial origin of clotting factor replacement cannot fully explain the alterations in lymphocyte subset distribution previously described in patients with hemophilia A.