Evaluation of biological specimen acceptability in a complex clinical laboratory before and after implementing automated grading serum indices.

Continuous monitoring of specimen acceptability, collection and transport can result in the prompt identification and correction of problems, leading to improved patient care and a reduction in unnecessary redraws and delays in reporting results. This study aims to identify unacceptable blood specimens and to calculate the specimen rejection rate (SRR) before and after the implementation of automated checks of serum indices. This study was conducted between January 2009 and December 2010. The number of rejected specimens, location and reason for rejection were recorded. The Architect c8000 analyser (Abbott, Illinois, USA) was used to assess serum indices based on characteristic spectral patterns and mathematical manipulations of absorbance values measured at several wavelengths. The SRR was calculated, and the target cut-off value for the SRR was < 0.5%, as established by the College of American Pathologists (CAP). The SRR values were 0.13% and 0.21% for the years 2009 and 2010, respectively. Haemolysis was the most significant reason for sample rejection, with cumulative rejection rates (CRR) of 49.3% and 61.4% for 2009 and 2010, respectively. Adult intensive care units (ICUs) had the most sample rejections (23.5%), followed by neonatal ICUs (13.8%), cardiac ICUs (13%), paediatric ICUs (10.8%) and long-term wards (10.5%), of which 60%, 79%, 84.9%, 36.6% and 75%, respectively, of the rejected samples were haemolysed. The increase in rejected samples may be due to an improvement in staff awareness of sample rejection, aided by automatic sample integrity grading by automated chemistry analysis systems.

Thymoquinone, the active ingredient of Nigella sativa seeds, enhances survival and activity of antigen-specific CD8-positive T cells in vitro.

Recent preclinical and clinical studies provide evidence that adoptive transfer of in vitro activated T cells can results in significant antitumour responses in vivo upon acquisition of certain survival and homing properties during in vitro activation. Based on recent studies showing in vivo antioxidant effects of thymoquinone (TQ), the active ingredient of Nigella sativa seeds, this study aims to determine whether or not TQ can increase survival and sustain the expression of the homing receptor CD62L in antigen-specific T cells in vitro. The results showed that stimulation of OT-1 (transgenic CD+) T cells with OVA antigen resulted in activation, as shown by a decrease in the surface expression of CD62L which coincided with significant apoptosis measured three and five days after antigen stimulation. Addition of low concentrations of TQ during CD85+ T-cell activation resulted in enhanced survival of the activated T cells and sustained expression of CD62L. These effects coincided with enhancement in the capability of CD8+ T cells to produce the effector cytokine interferon-gamma (IFNgamma). These results suggest that TQ has a beneficial effect in conditioning T cells in vitro for adoptive T-cell therapy against cancer and infectious disease.

The use of growth factors to modulate the activities of antigen-specific CD8+ T cells in vitro.

RATIONALE: Adoptive T cell therapy depends on the harvesting of the cells from the host, their activation in vitro, and their infusion back to the same host. The way of activating the T cells in vitro is a critical factor for their homing, survival and function in vivo. Sustaining T cell homing molecules, particularly CD62L, is benefic for the trafficking of the adoptive transferred cells. OBJECTIVE: The aim of the present study is to test whether insulin-like growth factor-1 (IGF-1), thymosin- α1 (T-α1) as well as all-trans retinoid acid (ATRA) alone or in combination with IL-2, IL-12, IL-15 can enhance the activation and survival phenotypes of antigen-activated T cells in vitro. METHODS & RESULTS: To this end, OT-1 transgenic T cells were used as a model. These CD8+ T cells recognize OVA peptide presented by MHC class-I. The results showed that antigen stimulation of OT1 cells resulted in their activation as evidenced by the decrease in surface expression of CD62L, analyzed for 3 days after antigen stimulation and was more pronounced on day 5. The addition of IL-12 or IGF-1 alone but not of IL-2, IL-15 augmented OT-1 cell activation measured on day 5. Interestingly, the combination of IL-12 with IGF-1 sustained the expression of CD62L on OT1 cells. Although the addition of ATRA alone or in combination with IL-12 resulted in decreases in CD62L expression on day 3, they showed a dose-dependent effect on the restoration of CD62L expression on day 5. The analysis of the activation-induced cell death (apoptosis) of OT1 cells showed an increased rate of death on day 5 than on day 3-post antigen stimulation. The addition of only IL-12 or IGF-1 alone, but not of IL-2, IL-15 or T- α1, decreased OT1 cell apoptosis on day 3. These anti-apoptotic effects of IL-12 and IGF- 1, however, were recovered on day 5-post stimulation. DISCUSSION: In conclusion, these results indicate that the activation phenotype and the survival of antigen-specific T cells can be differently modulated by immunomodulatory factors, where, interleukin-12 and IGF-1 induced the favorable effect. These results have a significant implication for T cell adoptive immunotherapy in different settings.

Protective effects of Nigella sativa oil and thymoquinone against toxicity induced by the anticancer drug cyclophosphamide.

Constituents of the Nigella sativa seed are reported to possess potent antioxidant effects. Treatment with anticancer drugs such as cyclophosphamide (CTX) is associated with significant toxicity due to over-production of reactive oxygen species, resulting in increased levels of oxidative stress. The aim of this study is to test whether or not N. sativa L oil (NSO) or its active ingredient, thymoquinone (TQ), can reduce CTX-induced toxicity. Male albino rats were treated with intraperitoneal administration of phosphate buffered saline (PBS) or 200 mg/Kg CTX followed by intragastric administration of NSO or TQ on alternate days for 12 days. Administration of NSO and TQ was initiated 6 h before or after CTX injection. Twenty-four hours after the last NSO and TQ treatment, blood and liver were harvested to analyse toxicity-related parameters. Treatment with CTX induced significant toxicity as shown by decrease in haemoglobin concentration and increases in blood sugar levels, activities of liver enzymes, bilirubin, urea, creatinine, lipids (triglyceride, cholesterol and low-density lipoprotein (LDL)-cholesterol) and lipid peroxidation in the liver. Treatment with NSO or TQ induced significant reduction in overall toxicity. The antitoxic effects of NSO and TQ were associated with induction of antioxidant mechanisms. These results suggest that administration of NSO or TQ can lower CTX-induced toxicity as shown by an up-regulation of antioxidant mechanisms, indicating a potential clinical application for these agents to minimise the toxic effects of treatment with anticancer drugs.

Vaccines, virucides and drugs against HIV/AIDS: hopes and optimisms for the future.

More than 25 million lives have been claimed by AIDS and 33.2 million people are estimated to have HIV, the majority of which are living in the underdeveloped countries. Failed tests on vaccines, virucides and complete virus eradication have caused scientists to refocus on the basic questions of what makes an effective HIV immune response. The "gloom" over disappointing research results on vaccine development and virucides "threatens to overshadow more positive" HIV/AIDS-related news, such as findings that male circumcision might reduce the likelihood of HIV transmission and that giving antiretroviral drugs to "high-risk" HIV-negative people (pre-exposure prophylaxis) could help protect them from infection. Something like pre-exposure prophylaxis has a good chance of becoming available before we have a 100% efficacious vaccine. The future in the field of HIV/AIDS will be much brighter if global research is appropriately coordinated and sufficient funds are available.

Links between apoptosis, proliferation and the cell cycle.

Many physiological processes, including proper tissue development and homeostasis, require a balance between apoptosis and cell proliferation. All somatic cells proliferate via a mitotic process determined by progression through the cell cycle. Apoptosis (programmed cell death) occurs in a wide variety of physiological settings, where its role is to remove harmful, damaged or unwanted cells. Apoptosis and cell proliferation are linked by cell-cycle regulators and apoptotic stimuli that affect both processes. This review covers recent developments in the field and examines new evidence of the interconnection between apoptosis and cell proliferation.

Induction of apoptosis in myeloid progenitors by granulocyte-macrophage colony-stimulating factor.

Spontaneous apoptosis of normal purified bone marrow CD34+ cells induced by granulocyte-macrophage colony-stimulating factor (GM-CSF) via the Fas pathway appears to be mediated by caspase-1 and caspase-8 activity. In seeking an alternative explanation for this observation, the present study examined CD34+ cell growth with different cytokines, cytokine concentrations, caspase inhibitors, cell crowding and different media. Exposure of the normal CD34+ cells to different concentrations of GM-CSF and granulocyte colony-stimulating factor (G-CSF) increased apoptosis at lower concentrations. However, these GM-CSF effects were suppressed by G-CSF. Investigation of the association between apoptosis and crowding and different media showed that: 1) G-CSF and GM-CSF are equally effective as survival factors, and 2) the percentage of apoptotic cells in liquid culture was markedly lower than that found in methylcellulose culture. Finally, immunofluorescence staining showed that Fas was expressed at 10 ng/mL GM-CSF, while Bcl-2 expression was detected at 100 ng/mL. These findings suggest that cytokine concentration, cell culture conditions, cell crowding and cell interactions all are important factors in GM-CSF-induced apoptosis.

Impact of new AABB guidelines on hepatitis B and C testing among Saudi blood donors.

This study aims to compare the advantages and disadvantages of implementing the new American Association of Blood Banks (AABB) guidelines for hepatitis B and C against its old criteria for screening blood donors. Between July 1995 and December 2002, 63,368 consecutive blood donors were screening for hepatitis B and C according to the new guidelines. Cost and prevalence were analysed and compared with those found using the old AABB guidelines prior to July 1995. The overall percentage rate of deferred donors showed a significantly decrease to 19.3% in 2002, compared to 58.4% before July 1995 (P < 0.001). The new prevalence of hepatitis B and C among Saudi blood donors was found to be 1.7% and 0.6%, respectively, compared to 4% and 1.4%, respectively, under the old AABB guidelines. This resulted in a significant increase in the number and yield of blood units, and a decrease in the prevalence of hepatitis B and C was observed among screened donors. Using the new AABB guidelines, the estimated direct cost of donor screening for hepatitis B and C decreased significantly from 42.8 dollars per donor to 29.2 dollars per donor (P<0.001).

Peripheral blood progenitor cell mobilisation alters myeloid, but not erythroid, progenitor cell self-renewal kinetics.

Transplantation of progenitor cells which have been mobilised into the bloodstream (PBPC) following the administration of G-CSF results in more rapid neutrophil recovery than transplantation of bone marrow (BM). The reasons for the accelerated neutrophil engraftment are not clear, but would be explained by increased self-replication of myeloid progenitor cells (CFU-GM). We have used a CFU-GM replating assay to investigate myeloid progenitor self-replication, and quantification of subcolony formation during erythroid burst formation to quantify erythroid progenitor self-renewal. Secondary colony formation by CFU-GM, grown from PBPC and then replated was increased compared with secondary colony formation by BM CFU-GM (P = 0.0001); erythroid subcolony formation was not altered. There was no difference between the replating abilities of PBPC CFU-GM derived from allogeneic donors (normal individuals) and autologous donors (patients with malignant disease) although differences were found between subgroups of autologous donors. The increased replication of PBPC could not be accounted for by a reduction in progenitor cell apoptosis; PBPC CFU-GM contained slightly fewer apoptotic CD34+ cells than BM CFU-GM. The increased replication by PBPC CFU-GM was reversible because it declined when CFU-GM colonies were passaged through three sequential CFU-GM replating cycles. This decline in self-replication was more rapid than the decline seen in replated BM CFU-GM. The self-replication of PBPC CFU-GM, and subcolony formation by BFU-E could be further enhanced by exposure to cytokines in vitro. We conclude that mobilisation alters the replication kinetics of myeloid, but not of erythroid, progenitor cells, that mobilisation-induced events are of limited duration and that in vitro exposure to cytokines may modify PBPC progenitor cell kinetics.