Two doses of humanized anti-CD25 antibody in renal transplantation: A preliminary comparative study

HuCD25mAb is a humanized anti-CD25 antibody which has the same amino acid sequence as daclizumab (Zenapax, Roche). HuCD25mAb is expressed in Chinese hamster ovary (CHO) cells while daclizumab is expressed in the NSO myeloma cell line. A comparative study was performed to evaluate the pharmacokinetics and pharmacodynamics between huCD25mAb and daclizumab in a two-dose regimen incorporating triple immunosuppressant treatment regimens (MMF, CsA and steroids). Fifteen patients were enrolled and randomized to receive intravenous infusion of either huCD25mAb (n = 10) or daclizumab (n = 5) at a dosage of 1 mg·kg⁻¹ on operation day 0 and post-operation day 14. Serum concentrations of huCD25mAb and daclizumab were measured by a validated competitive ELISA. Subgroups of CD3⁺, CD25⁺, CD4⁺ and CD8⁺ lymphocytes were monitored periodically by flow cytometry. The concentration-time curves of huCD25mAb and daclizumab were found to fit well to a one-compartment model. A significant decline of proportion (%) of CD3-CD25⁺ and CD3⁺CD25⁺ lymphocytes was observed 30 min after first infusion on day 0 (3.40 ± 1.83 to 0.03 ± 0.07, 3.35 ± 2.02 to 0.37 ± 0.49), and these levels remained low for at least 70 days (0.03 ± 0.05, 0.31 ± 0.47). All pharmacokinetic parameters of huCD25mAb seemed similar to those of daclizumab. The two-dose huCD25mAb regimen was as effective as daclizumab in rapidly achieving high therapeutic concentration in the treated patients, and a significant decrease of CD3⁻CD25⁺ and CD3⁺CD25⁺ lymphocytes was demonstrated. This suggests that two-dose regimen is feasible in maintaining host immunosuppression and may provide an effective and economical strategy for reducing incidence of acute graft rejection.Key words: CD25, pharmacokinetics, kidney transplantation, enzyme immunoassay, flow cytometry, monoclonal antibody     

Assays for mammalian tyrosinase: a comparative study

This work describes a comparative study of the tyrosinase activity determined using three methods which are the most extensively employed; two radiometric assays using L-tyrosine as substrate (tyrosine hydroxylase and melanin formation activities) and one spectrophotometric assay using L-dopa (dopa oxidase activity). The three methods were simultaneously employed to measure the activities of the soluble, melanosomal, and microsomal tyrosinase isozymes from Harding-Passey mouse melanoma through their purification processes. The aim of this study was to find any correlation among the tyrosinase activities measured by the three different assays and to determine whether that correlation varied with the isozyme and its degree of purification. The results show that mammalian tyrosinase has a greater turnover number for L-dopa than for L-tyrosine. Thus, enzyme activity, expressed as mumol of substrate transformed per min, is higher in assays using L-dopa as substrate than those using L-tyrosine. Moreover, the percentage of hydroxylated L-tyrosine that is converted into melanin is low and is affected by several factors, apparently decreasing the tyrosinase activity measured by the melanin formation assay. Bearing these considerations in mind, average interassay factors are proposed. Their values are 10 to transform melanin formation into tyrosine hydroxylase activity, 100 to transform tyrosine hydroxylase into dopa oxidase activity, and 1,000 to transform melanin formation into dopa oxidase activity. Variations in these values due to the presence in the tyrosinase preparations of either inhibitors or regulatory factors in melanogenesis independent of tyrosinase are also discussed.     

Homologous transplantation with fresh frozen bone for dental implant placement can induce HLA sensitization: a preliminary study

It has been related in orthopedic surgeries the HLA sensitization. Thus, we evaluate if the use of fresh-frozen homologous bone (FFHB) for dental implant placement induce anti-HLA sensitization. Six patients were treated with FFHB corticocancellous block grafts. After 6 months, bone biopsies were harvested during implant placement to allow histomorphometric analysis. Vital mineralized tissue (VMT), non-vital mineralized tissue (NVMT) and non-mineralized tissue (NMT) were quantified histomorphometrically. Peripheral blood was collected from the patients before FFHB placement and 6 months after the surgery for anti-HLA analysis. The histomorphometric analysis showed the presence of VMT, NVMT and NMT in 45.56 ± 15.72 %, 14.16 ± 13.39 % and 40.29 ± 12.60 %, respectively. The baseline and 6 months postoperative CTs revealed bone thickness in the order of 5.66 ± 0.67 mm and 8.71 ± 1.52 mm (3.05 ± 1.39 mm). The anti-HLA analysis revealed that two of the six patients (33.3 %) became sensitized, however this was not associated with any FFHB incorporation loss (p > 0.05). A total of 24 implants were placed all of which were osseointegrated after 6 months. Although FFHB-related HLA sensitization does not appear to affect bone incorporation when treating insufficient bone thickness for implant placement, further follow-up is required to determine whether there is an association between HLA sensitization and long-term graft survival.     

Cell transplantation for treatment of acute myocardial infarction: unique capacity for repair by skeletal muscle satellite cells

An adult heart injured by an ischemic episode has a limited capacity to regenerate. We administered three types of adult guinea pig cells [cardiomyocytes (CMs), cardiac fibroblasts (CFs), and skeletal myoblasts (Mbs)] to compare their suitability for repair of acute myocardial infarction. We used confocal fluorescent microscopy and a variety of specific immunomarkers and echocardiography to provide anatomic evidence for the viability of such cells and their possible functional beneficial effects. All cells were transfected with adenovirus-containing beta-galactosidase gene so that migration from the injection sites could be traced. Both freshly isolated CMs as well as CFs were found concentrated in the infarcted zone; these cells survived for at least 2 wk posttransplantation. Transplanted CMs were regularly striated and grew long projections that could form gap junctions with native CMs, which was evidenced by connexin43 labeling. In addition, CM transplantation resulted in increased angiogenesis in the infarcted areas. In contrast, transplanted CFs did not appear to make any gap junctional contacts with native CMs nor did they enhance local angiogenesis. Mbs cultured for 7 days and transfected Mbs were identified 7 days posttransplantation in the infarcted area. During that time and thereafter, Mbs proliferated and differentiated into myotubes that formed new, regularly striated myofibers that occupied most (50-70%) of the infarcted area by 2-3 wk. These newly formed myofibers maintained their Mb skeletal muscle origin as evidenced by their capacity to express myogenin and fast skeletal myosin. This skeletal phenotype appeared to downregulate with time, and Mbs partially transdifferentiated into a cardiac phenotype as indicated by labeling for cardiac-specific troponin T and cardiac myosin heavy chain. By the third week posttransplantation, new myofibers formed apparent contacts with the native CMs via putative gap junctions that expressed connexin43. Myocardial performance of animals that were successfully transplanted with Mbs was improved.     

Cholangiocytes: Cell transplantation

Background

Due to significant limitations to the access to orthotropic liver transplantation, cell therapies for liver diseases have gained large interest worldwide.

Cell transplantation for diabetes

For 30 years there has been experimental work aimed at transplanting islets for the treatment of diabetes with a view to curing the disease and preventing the secondary complications. Many technical difficulties were experienced, first in isolating the islets without damaging them, and second in finding a suitable place to inject them, but until recently the results of a vascularized pancreas transplant have been superior to islet transplantation. In 2000, the group in Edmonton, headed by Shapiro, published encouraging results using a different immunosuppression in transplanting patients earlier in the course of their disease than had been attempted previously. The results were excellent at a year and good at 2 years in patients with Type I diabetes, however there was the rather worrying attrition at five years. Nevertheless, the Edmonton observations were proof of concept and have intensified interest in treating diabetes and other diseases where a specific protein synthesis was required by cell transplantation and/or genetic engineering. The recent interest in embryonic stem cells extenuated these efforts and progress is being made in defining the difficulties, which are greater than most workers would have predicted.In this review, the subject is discussed explaining where progress needs to be made in order to provide treatment that would be of value to patients.     

Ischemic myocardial injury in cultured heart cells: preliminary observations on morphology and beating activity

An in vitro model of myocardial ischemia has been established with primary monolayer cultures of neonatal rat heart cells. Ischemic conditions were simulated in vitro by subjecting the heart cell cultures to various levels of oxygen and glucose deprivation. After the ischemic treatments, cultures of beating muscle (M) cells were evaluated for functional and morphological changes. The experimental protocol consisted of treatment with 20% or 0% O2 and 1000, 500 or 0 mg glucose per 1 of medium for 4, 12 or 24 hr. Control cultures were treated with 20% O2 and 1000 mg glucose. The morphological alterations induced by the deficiency of O2 and glucose in the medium were the formation of pseudopodia and cytoplasmic vacuoles; increased cytoplasmic granulation; and the formation of abnormal cell shapes, such as long, spindly shaped M cells. There was a time-dependent decrease in beating activity as the M cells were exposed to longer durations of ischemic conditions. However, if the cultures were replenished with complete medium (1000 mg glucose) and 20% O2, the cells regained their ability to beat.     

Polymorphisms in glutathione S-transferase are risk factors for perioperative acute myocardial infarction after cardiac surgery: a preliminary study

In the present study we explored glutathione S-transferase (GST) polymorphisms in selected patients who experienced accelerated myocardial injury following open heart surgery and compared these to a control group of patients without postoperative complications. 758 Patients were enrolled from which 132 patients were selected to genotype analysis according to exclusion criteria. Patients were divided into the following groups: Group I: control patients (n = 78) without and Group II.: study patients (n = 54) with evidence of perioperative myocardial infarction. Genotyping for GSTP1 A (Ile105Ile/Ala113Ala), B (Ile105Val/Ala113Ala) and C (Ile105Val/Ala113Val) alleles was performed by using real-time-PCR. The heterozygous AC allele was nearly three times elevated (18.5 vs. 7.7 %) in the patients who suffered postoperative myocardial infarction compared to controls. Contrary, we found allele frequency of 14.1 % for homozygous BB allele in the control group whereas no such allele combination was present in the study group. These preliminary results may suggest the protective role for the B and C alleles during myocardial oxidative stress whereas the A allele may represent predisposing risk for cellular injury in patients undergoing cardiac surgery.     

Breast compression: a preliminary study

X-Ray mammography is the technique employed to image breast tumours; it is the gold standard for both diagnosis and screening¹. To increase the sensitivity of the test and to reduce the dose of X-radiation, breast compression is used. The effect of squeezing a simple model of a breast in which there is an idealized spherical tumour volume is investigated; the cases where the lesion has both a smaller, and a larger compressibility than the surrounding tissues are considered. Surface strain is computed as a function of the deformation of the lesion from its spherical shape. Evidence from earlier studies is adduced to show that when the lesion is malignant care should be exercised to avoid the possibility of disseminating cancer cells by the application of compressive forces.     

Patterns of fractures in accidental and non-accidental injury in children: a comparative study.

The incidence and pattern of fractures in children who had been abused were compared with those of fractures sustained by children of similar ages in whom abuse had been excluded. From 1976 to 1982 there were 35 children with fractures resulting from child abuse, and all were aged under 5. Of the 826 children in the control group, seen from January to June 1981, 85% were aged over 5. Abused children were much more likely to have multiple fractures (p less than 0.001) and bruising of the head and neck (p less than 0.001). Fractures of the ribs were common in children who had been abused, and their presence, in the absence of major chest trauma, strongly suggested that abuse was occurring. Injuries to the long bones were invariably spiral or oblique fractures or subperiosteal new bone formation--both "gripping or twisting" injuries. Spiral fracture of the humeral shaft was significantly more common (p less than 0.001) in the group of abused children. Classic metaphyseal chip fractures were uncommon. One child in eight aged under 18 months who sustains a fracture may be a victim of child abuse.     

Rapid screening: a comparative study

Although rapid screening of negative and inadequate cervical smears is a quality assurance requirement for all UK laboratories, there has been little attempt to standardize the method and laboratories make use of a number of different techniques and times. The aim of this study was to assess the sensitivity of these various techniques by measuring their ability to pick out known false-negative smears. Completed questionnaires from 123 laboratories across England revealed that 52% of laboratories use a "step" technique, 19% use "turret", 15% use random paths and 34% attempt to rescreen the whole slide quickly. Twenty-two percent of laboratories use a mixture of techniques. Timings are also variable, with the majority of laboratories allowing screeners to review slides at a pace decided by themselves but usually between 1 and 2 min. The study involved 120 participants who performed a total of 24 000 rapid screens. The results showed that, of the 90 abnormal slides used in the study, 62 cases (69%) were identified as abnormal or needing review by more than 50% of participants. Overall rapid screening picked out 58% of high-grade squamous abnormalities, 59% of low-grade abnormalities and 72% of glandular lesions. Step screening performed best, followed by whole slide/random and then turret. One minute was the optimum time and there was a significant fall in performance once individuals attempted to rescreen large numbers (>50). The most significant finding was the marked variation in the performance of individuals using the same slide sets.     

Cell therapy for myocardial infarction

The review discusses cell therapy; one of the most promising approaches to myocardial infarction treatment. The possibility to use cell material of various origins is analyzed. The review sums up data on the application of fetal and neonatal cardiomyocytes, myoblasts, bone marrow mononuclear fraction, hematopoietic and mesenchymal stem cells (MSX) as cell therapy agents. The conclusion is made that MSC are promising cell material for myocardial infarction therapy. MSC are able to migrate to the injured area, differentiate into myocardial lineage. They produce a wide range of factors that stimulate angiogenesis and increase viability of cells, including cardiomyocytes.     

Cell transplantation therapies for spinal cord injury focusing on bone marrow mesenchymal stem cells: Advances and challenges

Spinal cord injury (SCI) is a devastating condition with complex pathological mechanisms that lead to sensory, motor, and autonomic dysfunction below the site of injury. To date, no effective therapy is available for the treatment of SCI. Recently, bone marrow-derived mesenchymal stem cells (BMMSCs) have been considered to be the most promising source for cellular therapies following SCI. The objective of the present review is to summarize the most recent insights into the cellular and molecular mechanism using BMMSC therapy to treat SCI. In this work, we review the specific mechanism of BMMSCs in SCI repair mainly from the following aspects: Neuroprotection, axon sprouting and/or regeneration, myelin regeneration, inhibitory microenvironments, glial scar formation, immunomodulation, and angiogenesis. Additionally, we summarize the latest evidence on the application of BMMSCs in clinical trials and further discuss the challenges and future directions for stem cell therapy in SCI models.     

Sheep lymphocyte antigens: a preliminary study

Sera from 287 sheep were screened for cytotoxic antibodies against sheep lymphocytes. Forty four antisera were selected which provisionally define 13 lymphocyte antigens. The frequency of these antigens was studied in 305 sheep from 8 flocks of different breeds. Family studies confirm that inheritance of sheep lymphocyte antigens is controlled by the autosomal codominant genes of at least 2 linked loci.     

Bayesian meta-analysis models for microarray data: a comparative study

Background

With the growing abundance of microarray data, statistical methods are increasingly needed to integrate results across studies. Two common approaches for meta-analysis of microarrays include either combining gene expression measures across studies or combining summaries such as p-values, probabilities or ranks. Here, we compare two Bayesian meta-analysis models that are analogous to these methods.

Results

Two Bayesian meta-analysis models for microarray data have recently been introduced. The first model combines standardized gene expression measures across studies into an overall mean, accounting for inter-study variability, while the second combines probabilities of differential expression without combining expression values. Both models produce the gene-specific posterior probability of differential expression, which is the basis for inference. Since the standardized expression integration model includes inter-study variability, it may improve accuracy of results versus the probability integration model. However, due to the small number of studies typical in microarray meta-analyses, the variability between studies is challenging to estimate. The probability integration model eliminates the need to model variability between studies, and thus its implementation is more straightforward. We found in simulations of two and five studies that combining probabilities outperformed combining standardized gene expression measures for three comparison values: the percent of true discovered genes in meta-analysis versus individual studies; the percent of true genes omitted in meta-analysis versus separate studies, and the number of true discovered genes for fixed levels of Bayesian false discovery. We identified similar results when pooling two independent studies of Bacillus subtilis. We assumed that each study was produced from the same microarray platform with only two conditions: a treatment and control, and that the data sets were pre-scaled.

Conclusion

The Bayesian meta-analysis model that combines probabilities across studies does not aggregate gene expression measures, thus an inter-study variability parameter is not included in the model. This results in a simpler modeling approach than aggregating expression measures, which accounts for variability across studies. The probability integration model identified more true discovered genes and fewer true omitted genes than combining expression measures, for our data sets.     

Cardiac contraction motion compensation in gated myocardial perfusion SPECT: A comparative study

IntroductionCardiac contraction significantly degrades quality and quantitative accuracy of gated myocardial perfusion SPECT (MPS) images. In this study, we aimed to explore different techniques in motion-compensated temporal processing of MPS images and their impact on image quality and quantitative accuracy.Material and method50 patients without known heart condition underwent gated MPS. 3D motion compensation methods using Motion Freezing by Cedars Sinai (MF), Log-domain Diffeomorphic Demons (LDD) and Free-Form Deformation (FFD) were applied to warp all image phases to fit the end-diastolic (ED) phase. Afterwards, myocardial wall thickness, myocardial to blood pool contrast, and image contrast-to noise ratio (CNR) were measured in summed images with no motion compensation (NoMC) and compensated images (MF, LDD and FFD). Total Perfusion Defect (TPD) was derived from Cedars-Sinai software, on the basis of sex-specific normal limits.ResultLeft ventricle (LV) lateral wall thickness was reduced after applying motion compensation (p < 0.05). Myocardial to blood pool contrast and CNR in compensated images were greater than NoMC (p < 0.05). TPD_LDD was in good agreement with the corresponding TPD_MF (p = 0.13).ConclusionAll methods have improved image quality and quantitative performance relative to NoMC. LDD and FFD are fully automatic and do not require any manual intervention, while MF is dependent on contour definition. In terms of diagnostic parameters LDD is in good agreement with MF which is a clinically accepted method. Further investigation along with diagnostic reference standards, in order to specify diagnostic value of each technique is recommended.     

Expression of uncoupling protein 3 in mitochondria protects against stress-induced myocardial injury: a proteomic study

It has been confirmed that stress plays an important role in the induction and development of cardiovascular diseases, but its mechanism and molecular basis remain unknown. In the present study, a myocardial injury model induced by restraint stress was established in rat. To screen for the related proteins involved in stress-induced myocardial injury, proteomic techniques based on 2-DE and mass spectrometry were used. In our results, ten proteins were found to be altered. The expression of eight of these proteins was increased after restraint stress, including cardiac myosin heavy chain, dihydrolipoamide succinyltransferase component of 2-oxoglutarate dehydrogenase complex, mitochondrial aldehyde dehydrogenase, H⁺-transporting ATP synthase, albumin, and apolipoprotein A-I precursor. The expression of uncoupling protein 3 (UCP3) and mitochondrial aconitase was decreased. Most of the proteins were related to energy metabolism. Further research indicated that UCP3 may mediate the myocardial cell response induced by restraint stress.     

The sarcoplasmic reticulum: a comparative study

The diversity of cellular membrane structures associated with regulation of intracellular calcium level is described in several different groups of organisms and cells. All the instances reported refer to cellular processes related to movement, in which calcium ion acts as trigger and/or modulator. In addition, a simplified five-stage picture of the underlying view of evolution of these structures is presented. In short: the choice made by nature in using calcium as intracellular messenger was very early in the history of life; all cellular structures devoted to intracellular calcium regulation, from the simplest form of amoeba to the highly sophisticated apparatus of mammalian skeletal muscle, can be linked together in the chain of evolution. Because the evidence is still sparse, any conclusion more positive would be speculative and of little value. Hopefully, in the coming years, with a better understanding of membrane architecture as a whole and its protein components (i.e. calcium channels, calcium-binding proteins), we will be able to test the first segments of this evolutionary hypothesis.