Factors influencing post-cryopreserved CD34+ cells viability in the harvested products of autologous haematopoietic stem cells

The cryopreservation process of stem cells potentially cause the loss of CD34+ cells. The aim of this study is to evaluate association of patient, graft and technical characteristics with post cryopreserved CD34+ cells viability among lymphoproliferative disease namely multiple myeloma (MM) and lymphoma patients at Hospital Universiti Sains Malaysia (USM). This retrospective study was conducted in the Transplant Unit. A search of the hospital data (2008–2018) to identify 132 patients for both MM and lymphoma who underwent autologous peripheral blood haematopoietic stem cells (APBSC) mobilisation, and were successfully harvested and cryopreserved. Selected patients’ profile as well as selected parameters of stem cell mobilization and cryopreservation were obtained from laboratory information system (LIS), record unit and the Transplant Unit. Multiple logistic regression (MLR) was used to find significant associated factors and P < 0.05 was considered significant. The mean age of the patients was 39 years old with almost equal gender distribution and majority were lymphoma patients, 96 (72.7%) while 36 (27.3%) were multiple myeloma (MM) patients. The significant influencing factors of post-cryopreserved CD34+ cells viability were pre-cryopreserved CD34+ cell viability, total nucleated cells (TNC), and anti-platelet and antibiotics usage. Patients who are not on anti-platelet and have higher pre-cryopreserved CD34+ cells viability have higher chance for good post-cryopreserved CD34+ cells viability. While, those patients with higher TNC and on antibiotics have lower chance for good post cryopreserved CD34+ cells viability. This study showed patients who are not on anti-platelet and antibiotics will have higher probability of achieving good post cryopreserved CD34+ cells viability. The APBSC products with higher pre-cryopreserved CD34+ cells viability and lower TNC will achieve better post-cryopreserved CD34+ cells viability. The addition of extra plasma to the APBSC products is recommended to reduce the TNC.     

Novel colloidal associations of soyasaponins and lipid components (DPPC,cholesterol) as potential adjuvants for vaccines

Soyasaponins from soybean (Glycine max) represent promising new potent adjuvants for vaccine research because of their immunostimulating properties and weak hemolytic activity. In the present study, saponin microstructures of soyasaponins (soyasaponin Bb, soyasaponin Ab) with lipid components (cholesterol, DPPC (dipalmitoylphosphatidylcholine)) were designed by the lipid film method. In interaction studies between soyasaponins (soyasaponin Ab/Bb) and Langmuir monolayers (model membranes), composed of cholesterol and DPPC, marked interactions between soyasaponins and a pure cholesterol monolayer were observed. No interaction was detected for soyasaponins with a pure DPPC monolayer. The intercalation of soyasaponins in a mixed DPPC/cholesterol (3:1, w/w) monolayer was only observed for the monodesmosidic soyasaponin Bb whereas the second sugar chain of the bidesmosidic soyasaponin Ab impaired the access to the monolayer. Transmission electron microscopy was used for visualizing particle formation of soyasaponins and lipid components. Pseudo-binary systems (soyasaponin Ab/Bb, cholesterol) formed colloidal associations built up from ring-like subunits in the nanometer size range. In pseudo-ternary systems (soyasaponin, cholesterol, DPPC) soyasaponin Bb attacked the liposomal membrane by forming colloidal associations. Colloidal associations in pseudo-ternary systems with soyasaponin Ab, cholesterol and a phospholipid were only observed in the presence of PE (phosphatidylethanolamine) instead of DPPC. In an MTT assay with a HaCaT cell line (keratinocyte cell line) the cell viability was neither affected by the soyasaponins nor by the corresponding formulations. Both the pure soyasaponin solution and the saponin formulations may be promising adjuvant systems for the intradermal vaccine application. Furthermore, interaction studies between the model antigen ovalbumin and colloidal associations of saponins and cholesterol using MST (Microscale Thermophoresis) gave first indications of an antigen binding to colloidal associations. Ex vivo T-cell proliferation in the presence of soyasaponin Ab was confirmed.     

Viability evaluation of freeze dried and suspension anthrax spore vaccine formulations stored at different temperatures

Anthrax is endemic in Ethiopia with sporadic outbreaks despite the regular vaccination of domestic livestock. This has raised concerns on the effectiveness of the vaccination strategy which may be associated with breaches in the vaccine cold chain maintenance. This study was aimed at demonstrating the tolerance of anthrax vaccine to cold chain breaches through evaluation of viable spore counts expressed as colony forming units per mL (CFU/mL) of freeze-dried and suspension anthrax vaccines stored at 5 °C, 20 °C and 37 °C for up to 6 months. Both vaccine formulations maintained above the recommended minimum required titre (2 × 10⁶ culturable spores per dose for cattle, buffaloes and horses, and not <1 × 10⁶ for sheep and goats) for up to 6 months at 5 °C storage. In storage at 20 °C, the viability of freeze-dried anthrax vaccine maintained the minimum required titre up to 6 months while up to 90 days in case of the suspension formulation. Both types of vaccine formulations maintained the minimum titre per dose for up to 30 days at 37 °C storage. Generally, both vaccine formulations showed similar trends in titre fall in all of the three storage temperatures (5 °C, 20 °C and 37 °C) as observed in the almost linearly overlapping 95% confidence intervals (CI) up to day 90 at 5 °C and 20 °C storages while up to day 30 at 37 °C storage. However, a significant (P < 0.05) drop in titre was observed after day 90 for storages at 5 °C and 20 °C, and after day 30 for 37 °C storage as observed in the non overlapping 95% CI from the average titres of previous time points. This study showed that if temperature excursion occurs above the recommended temperature range (4–8 °C) during storage or transport, the vaccine should remain effective and can still be used in vaccination programs.     

Immunostimulatory Effects of ��-glucan Purified from Paenibacillus polymyxa JB115 on Mouse Splenocytes

We investigated the effects of β-glucan purified from Paenibacillus polymyxa JB115 on the viability and proliferation of splenocytes. Splenocytes play a critical role in host immunity. MTT assays and trypan blue exclusion tests revealed that β-glucan significantly promoted the viability and proliferation of splenocytes over a range of concentrations. However, there was no specific subset change. β-glucan protected splenocytes from cytokine withdrawal-induced spontaneous cell death. For further mechanistic studies, ELISA assay revealed that β-glucan enhanced the expression of anti-apoptotic molecules and interleukin 7 (IL-7), a cytokine critical for lymphocyte survival. We also investigated the IL-2 dependency of β-glucan-treated splenocytes to determine if treated cells could still undergo clonal expansion. In flow cytometric analysis, β-glucan induced increased levels of the activation marker CD25 on the surface of splenocytes and β-glucan-treated splenocytes showed higher proliferation rates in response to IL-2 treatment. This study demonstrates that β-glucan can enhance the survival of splenocytes and provides valuable information to broaden the use of β-glucan in research fields.     

Comparison of XTT and Alamar blue assays in the assessment of the viability of various human cancer cell lines by AT-101 (−/− gossypol)

This study compared the two different commercially available in vitro viability assays: XTT and Alamar blue (AB), to detect anti-proliferative effects of AT-101, a cotton plant extract, on six different human carcinoma cell lines including: prostate (PC-3 and DU-145), breast (MCF-7 and MDA-MB-231), and ovary (OVCAR-3 and MDAH 2774) in a time- and dose-dependent manner. Cells were exposed to AT-101 in the concentration range of 2.5–40 µM for 24, 48, and 72?h. The AB assay was slightly more sensitive than the XTT assay in the evaluation of AT-101 at 24?h, suggesting that the AB assay might be used for detecting early changes in cell viability as compared to the XTT assay. Moreover, the AB assay showed less intra-assay variability as compared to the XTT. The non-toxic, non-radioactive AB metabolism assay allows rapid assessment of large numbers of samples, with simple equipment and at reduced cost for continuous monitoring of cancer cell viability, and, thus, should be accepted as a suitable alternative viability method.     

湖北钉螺在非适宜地东平湖生存繁殖的纵向观察

目的在南水北调条件下,观察人工北移至非适宜地区山东省东平湖区湖北钉螺生存繁殖的能力。方法近年,在东平湖区的湖心岛以螺笼放养钉螺,定量观察其生存繁殖情况。结果亲代钉螺可以在东平湖越冬生存,存活率为60.51%(118/195);经过2个冬季24个月放养,繁殖产生了子2代钉螺228只,其螺口数量较亲代钉螺数量增长了14.00%;经过3个冬季36个月放养,钉螺繁殖产生了子3代钉螺86只,其数量较亲代钉螺减少了57.00%,与子2代钉螺数量比较减少了62.28%;经过4个冬季48个月放养,钉螺存活率为0。结论南水北调东线南端长江取水口地区的湖北钉螺在血吸虫病非流行区山东省东平湖区可以繁殖到子3代,生存繁殖的时间不超过4个冬季、48个月,呈现逐年消亡趋势。报道了在中国大陆北纬35 055/地区,人工北移到此地的钉螺在湖滩铁丝笼内放养状态下生存繁殖的纵向观察结果。