Immune regulation in plasma cell myeloma

University of Technology, Sydney. Faculty of Science.The pathogenesis of monoclonal gammopathies, particularly plasma cell myeloma (MM), is multifaceted and complex. In recent years, Treg and Th17 cells have emerged as key factors in the development and progression of malignancies including MM. However, there are still conflicting reports on whether regulatory T (Treg) cells and Th17 cells are increased or decreased in the peripheral blood (PB) of patients with MM. This is partly due to technical difficulties associated with the use of the transcriptional repressor, forkhead box P3 (FoxP3) to identify Treg cells. Studies have shown FoxP3 results to be dependent on the clones, fluorochromes attached and fixation/permeabilisation methods used. More recent studies have defined Treg cells as CD4⁺CD25ͪͥ cells which do not express CD127, an IL-7 receptor. This methodology was used to determine Treg cell number and develop assays for the assessment of Treg cell function. The study also extends to exploring Th17 cell number in plasma cell dyscrasias and the overall effect of the Treg and Th17 cell equilibrium on the survival of patients with MM. CD4⁺CD25ͪͥCD127⁻ expression was used to quantitate Treg cell numbers and an intracellular IL-17 assay on CD3⁺CD4⁺ cells was used for Th17 cell enumeration. Treg cell function was determined using carboxyfluorescein succinimidyl ester (CFSE) tracking of Treg depleted lymphocyte preparations stimulated by anti-CD3,CD2,CD28 beads at a 1:1 ratio for 4 days ± 1:1 fluorescence-activated cell sorted Treg cells. This functional assay was also used to investigate the effect of recombinant human (rh) TGF-β and rhIL-12 on Treg cells. The mean proportion of Treg cells in the CD4⁺ compartment of PB of patients with MM (n=32) was 8.9±0.6% and this was increased compared to the mean of the normal cohort (n=36) at 6.5±0.4% (p=0.009). However, no significant difference was observed between the frequency of PB Treg cells in the control group compared to patients with monoclonal gammopathy of undetermined significance (MGUS) (n=20) (mean=7.5±0.8%; P=0.24) and patients with Waldenström macroglobulinaemia (WM) (n=13) (mean=6.0±0.5%; P=0.48). Interestingly, a comparison of the absolute numbers exhibited different results. A significantly lower number of Treg cells was observed in patients with MM [(3.2±0.4) x10⁷/L; P<0.01] and WM [(3.0±0.6) x10⁷/L; P<0.01] compared to the control group [(6.4±0.7) x10⁷/L]. However, no significant difference was observed when comparing patients with MGUS [(4.3±0.8) x10⁷/L; P= 0.06] to the normal cohort. It was observed that a significantly higher proportion of PB Treg cells in patients with MM (85.9±1.8%; P<0.01) and WM (86.4±2.1%; P=0.02) to be of the CD45RO⁺ memory phenotype compared to the normal cohort (76.7±2.5%). However this was not observed in patients with MGUS (73.4±4.0%; P=0.47). In addition, the study revealed that PB Treg cell proportions were not influenced by MM stage. Thalidomide treated patients with MM appeared to have an increased PB Treg cell proportion, however only a small number of thalidomide treated patients were tested due to the use of thalidomide therapy being phased out and its replacement with lenalidomide. Treg cells from bone marrow (BM) were compared to matched PB samples from patients with MM, demonstrating a significantly greater proportion (p=0.02) of Treg cells in the CD4⁺ compartment of the BM (9.7±1.2%) compared to PB (6.7±1.4%). Regarding Th17 cells, a significant decrease (p=0.03) in the mean proportion and absolute number of Th17 cells was observed in the PB of patients with MM (n=22) compared with the controls (n=20) (0.7±0.1% and 2.0±0.6% respectively). However, the mean number of Th17 cells in patients with MGUS (2.2±0.6%; n=15) and WM (1.1±0.2%; n=12) was not significantly different from normal. No correlation was observed between Th17 cell number and MM staging or therapy. Additionally, the study explored the Treg/Th17 cell ratio in PB of patients with monoclonal gammopathies with comparison made to normal subjects. The mean Treg/Th17 cell ratio of patients with MM (16.1±2.4) was significantly higher (p=0.0002) than the healthy control group (6.6±1.0). The Treg/Th17 cell ratio of WM and MGUS patients was 7.0±1.0 and 4.9±0.5 respectively, neither of which were statistically different to the ratio of the normal controls. Most interestingly, patients who have survived with MM for 10 or more years possessed a Treg/Th17 cell ratio similar to the normal controls (7.04±2.47; p=0.84) and this was shown to affect overall survival. The data demonstrated that patients with MM observed to have a high Treg/Th17 cell ratio had an overall shorter survival compared to those whose Treg/Th17 cell ratio was lower (p<0.025). The suppressive capability of Treg cells from MM patients (n=15) was variable. The Treg cell function of patients treated with lenalidomide (n=5) was increased (mean=68%) compared to patients treated with thalidomide (n=5; mean=23%), Velcade (n=3; mean=12%), untreated patients (n=5; mean=36%) and normal controls (n=11; mean=31%). The suppression exerted upon the CD4⁺ T cell subset in patients treated with bortezomib was significantly lower when compared to the normal cohort. However, no significant difference in CD8⁺ T cell suppression was found between patients with MM and the normal controls. rhTGF-β increased the suppressive capabilities and rhIL-12 reduced the function of Treg cells from both MM and normal PB samples. In conclusion, immune regulation is dysfunctional in patients with MM as the proportion of PB Treg cells is increased and Th17 cells are reduced. Also, the cytokine microenvironment and treatment have a major impact on the function of Treg cells. The data clearly delineate the importance of the PB Treg/Th17 cell equilibrium, revealing a strong association between the Treg/Th17 cell homeostatic balance and disease progression and survival in MM, indicating an imbalance may cause either or both the innate and adaptive immune system to be dormant and incapacitate the anti-tumour response

Phenotypic and genotypic characterization of methicillin-resistant Staphylococcus aureus (MRSA) isolated from dogs and cats at University Veterinary Hospital, Universiti Putra Malaysia.

Methicillin-resistant Staphylococcus aureus (MRSA) is known to cause nosocomial infections and is now becoming an emerging problem in veterinary medicine. The objective of the study was to determine the presence of MRSA in 100 cats and dogs sampled between November 2007 and April 2008 at the University Veterinary Hospital, Universiti Putra Malaysia. MRSA was detected in 8% of pets sampled. Ten percent (5/50) and 6% (3/50) of the isolates were from dogs and cats, respectively. All MRSA isolates possessed the mecA gene and were found to be resistant to at least three antimicrobials with a minimum of Oxacillin MIC of 8 µg/mL. One isolate (CT04) had an extremely high MIC of >256 µg/mL. The MLST type ST59 found in this study have been reported earlier from Singapore and other countries as a strain from animal and community-associated MRSA respectively. Pulsed-field gel electrophoresis revealed five pulsotypes. Two isolates from cats (CT27 and CT33) and three isolates from dogs (DG16, DG20, and DG49) were respectively assigned to pulsotypes B and D. The study suggests that cats and dogs in Malaysia are potential reservoirs for MRSA

Made-to-measure malaria vector control strategies: rational design based on insecticide properties and coverage of blood resources for mosquitoes.

Eliminating malaria from highly endemic settings will require unprecedented levels of vector control. To suppress mosquito populations, vector control products targeting their blood hosts must attain high biological coverage of all available sources, rather than merely high demographic coverage of a targeted resource subset, such as humans while asleep indoors. Beyond defining biological coverage in a measurable way, the proportion of blood meals obtained from humans and the proportion of bites upon unprotected humans occurring indoors also suggest optimal target product profiles for delivering insecticides to humans or livestock. For vectors that feed only occasionally upon humans, preferred animal hosts may be optimal targets for mosquito-toxic insecticides, and vapour-phase insecticides optimized to maximize repellency, rather than toxicity, may be ideal for directly protecting people against indoor and outdoor exposure. However, for vectors that primarily feed upon people, repellent vapour-phase insecticides may be inferior to toxic ones and may undermine the impact of contact insecticides applied to human sleeping spaces, houses or clothing if combined in the same time and place. These concepts are also applicable to other mosquito-borne anthroponoses so that diverse target species could be simultaneously controlled with integrated vector management programmes. Measurements of these two crucial mosquito behavioural parameters should now be integrated into programmatically funded, longitudinal, national-scale entomological monitoring systems to inform selection of available technologies and investment in developing new ones

Long-term survival in multiple myeloma is associated with a distinct immunological profile, which includes proliferative cytotoxic T-cell clones and a favourable Treg/Th17 balance

Despite improved outcomes in multiple myeloma (MM), a cure remains elusive. However, even before the current therapeutic era, 5% of patients survived >10 years and we propose that immune factors contribute to this longer survival. We identified patient

Components of the ribosome biogenesis pathway underlie establishment of telomere length set point in Arabidopsis

Telomeres cap the physical ends of eukaryotic chromosomes to ensure complete DNA replication and genome stability. Heritable natural variation in telomere length exists in yeast, mice, plants and humans at birth; however, major effect loci underlying such polymorphism remain elusive. Here, we employ quantitative trait locus (QTL) mapping and transgenic manipulations to identify genes controlling telomere length set point in a multi-parent Arabidopsis thaliana mapping population. We detect several QTL explaining 63.7% of the total telomere length variation in the Arabidopsis MAGIC population. Loss-of-function mutants of the NOP2A candidate gene located inside the largest effect QTL and of two other ribosomal genes RPL5A and RPL5B establish a shorter telomere length set point than wild type. These findings indicate that evolutionarily conserved components of ribosome biogenesis and cell proliferation pathways promote telomere elongation

Evaluation of alternative mosquito sampling methods for malaria vectors in Lowland South - East Zambia.

Sampling malaria vectors and measuring their biting density is of paramount importance for entomological surveys of malaria transmission. Human landing catch (HLC) has been traditionally regarded as a gold standard method for surveying human exposure to mosquito bites. However, due to the risk of human participant exposure to mosquito-borne parasites and viruses, a variety of alternative, exposure-free trapping methods were compared in lowland, south-east Zambia. Centres for Disease Control and Prevention miniature light trap (CDC-LT), Ifakara Tent Trap model C (ITT-C), resting boxes (RB) and window exit traps (WET) were all compared with HLC using a 3 × 3 Latin Squares design replicated in 4 blocks of 3 houses with long lasting insecticidal nets, half of which were also sprayed with a residual deltamethrin formulation, which was repeated for 10 rounds of 3 nights of rotation each during both the dry and wet seasons. The mean catches of HLC indoor, HLC outdoor, CDC-LT, ITT-C, WET, RB indoor and RB outdoor, were 1.687, 1.004, 3.267, 0.088, 0.004, 0.000 and 0.008 for Anopheles quadriannulatus Theobald respectively, and 7.287, 6.784, 10.958, 5.875, 0.296, 0.158 and 0.458, for An. funestus Giles, respectively. Indoor CDC-LT was more efficient in sampling An. quadriannulatus and An. funestus than HLC indoor (Relative rate [95% Confidence Interval] = 1.873 [1.653, 2.122] and 1.532 [1.441, 1.628], respectively, P < 0.001 for both). ITT-C was the only other alternative which had comparable sensitivity (RR = 0.821 [0.765, 0.881], P < 0.001), relative to HLC indoor other than CDC-LT for sampling An. funestus. While the two most sensitive exposure-free techniques primarily capture host-seeking mosquitoes, both have substantial disadvantages for routine community-based surveillance applications: the CDC-LT requires regular recharging of batteries while the bulkiness of ITT-C makes it difficult to move between sampling locations. RB placed indoors or outdoors and WET had consistently poor sensitivity so it may be useful to evaluate additional alternative methods, such as pyrethrum spray catches and back packer aspirators, for catching resting mosquitoes

A Two-cohort Phase I Study of Weekly Oxaliplatin and Gemcitabine, Then Oxaliplatin, Gemcitabine, and Erlotinib During Radiotherapy for Unresectable Pancreatic Carcinoma

Gemcitabine is a potent radiosensitizer. When combined with standard radiotherapy (XRT) the gemcitabine dose must be reduced to about 10% of its conventional dose. Oxaliplatin and erlotinib also have radiosensitizing properties. In vitro, oxaliplatin and gemcitabine have demonstrated synergy. We aimed to determine the maximum tolerated dose of oxaliplatin and gemcitabine with concurrent XRT, then oxaliplatin, gemcitaibine and erlotinib with XRT in the treatment of locally advanced and low volume metastatic pancreatic or biliary cancer

Eliminating Malaria Vectors.

Malaria vectors which predominantly feed indoors upon humans have been locally eliminated from several settings with insecticide treated nets (ITNs), indoor residual spraying or larval source management. Recent dramatic declines of An. gambiae in east Africa with imperfect ITN coverage suggest mosquito populations can rapidly collapse when forced below realistically achievable, non-zero thresholds of density and supporting resource availability. Here we explain why insecticide-based mosquito elimination strategies are feasible, desirable and can be extended to a wider variety of species by expanding the vector control arsenal to cover a broader spectrum of the resources they need to survive. The greatest advantage of eliminating mosquitoes, rather than merely controlling them, is that this precludes local selection for behavioural or physiological resistance traits. The greatest challenges are therefore to achieve high biological coverage of targeted resources rapidly enough to prevent local emergence of resistance and to then continually exclude, monitor for and respond to re-invasion from external populations