Epstein-Barr Virus Status and Subsequent Thiopurine Exposure Within a Paediatric Inflammatory Bowel Disease Population.

ABSTRACT: The use of thiopurine therapy in Epstein-Barr virus (EBV)-naïve inflammatory bowel disease (IBD) patients remains controversial due to a risk of EBV-associated complications. We evaluated EBV status and outcomes within our paediatric IBD population over an 8-year period; finding that 217 of 409 (53%) screened patients were seropositive for EBV at IBD diagnosis; that thiopurines were used in 189 of 217 (87%) seropositive and 159 of 192 (83%) seronegative patients (P = 0.22); and that 7 of 192 (4%) previously seronegative patients subsequently tested positive for EBV with 6 of 7 (86%) patients having concurrently recorded thiopurine use. All six patients continued thiopurine with/without a period of cessation; no EBV-associated lymphoproliferative disorders/serious complications were recorded within our cohort. A significant proportion of our patients would not receive thiopurine therapy should their use be avoided in EBV-negative patients (47%) or seronegative males (30%). The small but significant risks of thiopurine treatment must be balanced against the potential benefits of successful IBD management; further research into this is required.

A Decade of Varicella Screening Within a Paediatric Inflammatory Bowel Disease Population.

INTRODUCTION: Increased risk of opportunistic infection-e.g., varicella zoster infection-secondary to therapies is a cause of morbidity in inflammatory bowel disease [IBD] patients. The UK vaccination schedule does not include varicella immunisation. We aimed to evaluate the varicella screening and immunisation programme in a paediatric IBD population. METHODS: Data regarding IBD diagnosis, varicella status, and consequent immunisations/treatment interventions were collected retrospectively from the records of patients diagnosed with IBD over a 10-year period [2009-2018]. RESULTS: In all, 520 IBD patients were diagnosed; 505/520 [97%] had varicella testing; 46/505 [9%] were naïve. Of 501 patients, 391[78%] were tested before or within 7 days of diagnosis; this increased in the second 5-year period compared with the first (229/268 [85%] versus 162/233 [70%]; p <0.00001). Median diagnosis age of naïve patients was lower [8.3 years versus 12.8 years; p <0.00001]. Where vaccination was feasible, 21/31 [68%] had two and 7/31 [23%] one immunisation. Prednisolone induction led to lower rates of vaccination (5/13 [39%] versus 23/33 [70%] for other induction therapies; p =0.02). Of 28 vaccinated patients, 5 [18%] had suspected breakthrough varicella; and 6/18 [33%] unimmunised patients required post-exposure prophylaxis or treatment for varicella. Immunisation was associated with a decrease in patients requiring post-exposure prophylaxis (0/28 [0%] versus 5/18 [28%]; p =0.0006) and varicella-related hospital admission (1/28 [4%] versus 4/18 [22%]; p =0.01). CONCLUSIONS: High rates of varicella screening and immunisation within a PIBD population are possible, resulting in a reduction in hospital admissions for varicella treatment. Varicella immunisation may be of increasing importance within the PIBD population with the emergence of novel therapeutic strategies.

Prophylactic low dose continuous calcium infusion during peripheral blood stem cell (PBSC) collections to reduce citrate related toxicity.

BACKGROUND: Citrate toxicity is one of the most frequent complications of apheresis procedures. It is caused by the infusion of the acid citrate dextrose (ACD), which chelates the calcium ions. AIMS: The aim of this study is to assess the effectiveness of prophylactic continuous infusion of calcium gluconate over intermittent bolus infusion to reduce citrate toxicity during large volume peripheral blood stem cell collection. MATERIALS AND METHODS: We retrospectively analysed the records of PBSC collection procedures performed from March 2010 to December 2013. Donors were selected as per the set guidelines. Machine used to perform the procedures was Cobe spectra. The study population was divided into 2 groups. One composed of intermittent intravenous bolus infusion at the onset of hypocalcaemic symptoms, the other composed of calcium gluconate administration as continuous infusion throughout the procedure. RESULT: The most common reported hypocalcaemic symptoms were mild perioral paresthesia followed by digital numbness. Of the 50 individuals who were injected with bolus calcium 40 (80%) individuals suffered from symptoms of hypocalcaemia, whereas 23 of 66 individuals (34.8%) suffered from hypocalcaemia in the continuous infusion group. This difference was significant (P < 0.001). Both groups were compared with respect to age, gender ratio, weight of the individuals, total blood volume processed, ACD used, calcium gluconate dose used, time taken for the procedure, the product volume. Significant difference was noticed only with respect to the product volume. This implies that the groups were comparable with respect to parameters such as age, gender ratio, weight of the individuals, total blood volume processed, ACD used, calcium gluconate dose used, and the time taken for the procedure. Also that significantly more products (244 v/s 204 ml) was collected in the continuous infusion group. CONCLUSIONS: Our results show that prophylactic continuous IV administration of low dose calcium-gluconate throughout the PBSC harvesting procedure reduced the incidence as well as the severity of citrate related toxicity. This increases his/her tolerance to withstand longer durations of the procedure and collect more volume of the product, hence may reduce the number of sittings of the procedure.

Seroprevalence of infectious markers & their trends in blood donors in a hospital based blood bank in north India.

BACKGROUND & OBJECTIVES: Hepatitis B virus (HBV), human immunodeficiency virus (HIV), hepatitis C virus (HCV) and syphilis infections pose a great threat to blood safety. This study was undertaken to investigate the seroprevalence of serologic markers for transfusion transmitted infections (TTIs) among blood donors at a hospital based blood centre in north India over a period of nine years. METHODS: The results of serologic markers for TTIs (HBsAg, anti-HCV, anti-HIV and syphilis) of all blood donations (both voluntary and replacement) at our hospital from January 2005 to December 2013 were screened. Additional analysis was conducted to examine the prevalence trends associated with each of the positive marker. RESULTS: The data of 180,477 donors [173,019 (95.86%) males and 7,458 (4.13%) females] were analyzed. Replacement donations [174,939 (96.93%)] represented the majority whereas, only 5,538 (3.06%) donations were from the voluntary donors. The risk of blood being reactive was three times higher in male donors when compared with the female donors. The risk of blood being reactive for one or more infectious markers was 2.1 times higher in replacement donors when compared with the voluntary donors. Seropositivity of HIV, HBsAg, HBcAb, syphilis showed a significant decreasing trend (P<0.05) while there was an increasing trend in HCV infection which was insignificant. INTERPRETATION & CONCLUSIONS: This study reflects that the risk of TTIs has been decreased over time with respect to HIV, HBV and syphilis, but the trends for HCV remains almost the same in blood donors. Blood transfusion remains a risk factor for the spread of blood-borne infections. Therefore, improvements are needed to strengthen both safety and availability of blood.

A multivariate analysis to assess the effect of packed red cell transfusion and the unit age of transfused red cells on postoperative complications in patients undergoing cardiac surgeries.

BACKGROUND: Transfusion of blood components and age of transfused packed red cells (PRCs) are independent risk factors for morbidity and mortality in cardiac surgeries. MATERIALS AND METHODS: We retrospectively examined data of patients undergoing cardiac surgery at our institute from January 1, 2012 to September 30, 2012. Details of transfusion (autologous and allogenic), postoperative length of stay (PLOS), postoperative complications were recorded along with other relevant details. The analysis was done in two stages, in the first both transfused and nontransfused individuals and in the second only transfused individuals were considered. Age of transfused red cells as a cause of morbidity was analyzed only in the second stage. RESULTS: Of the 762 patients included in the study, 613 (80.4%) were males and 149 (19.6%) were females. Multivariate analysis revealed that factors like the number and age of transfused PRCs and age of the patient had significant bearing upon the morbidity. Morbidity was significantly higher in the patients transfused with allogenic PRCs when compared with the patients not receiving any transfusion irrespective of the age of transfused PRCs. Transfusion of PRC of over 21 days was associated with higher postoperative complications, but not with in-hospital mortality. CONCLUSION: In patients undergoing cardiac surgery, allogenic blood transfusion increases morbidity. The age of PRCs transfused has a significant bearing on morbidity, but not on in-hospital mortality. Blood transfusion services will therefore have to weigh the risks and benefits of providing blood older than 21 days in cardiac surgeries.

In vivo gene silencing following non-invasive siRNA delivery into the skin using a novel topical formulation.

Therapeutics based on short interfering RNAs (siRNAs), which act by inhibiting the expression of target transcripts, represent a novel class of potent and highly specific next-generation treatments for human skin diseases. Unfortunately, the intrinsic barrier properties of the skin combined with the large size and negative charge of siRNAs make epidermal delivery of these macromolecules quite challenging. To help evaluate the in vivo activity of these therapeutics and refine delivery strategies we generated an innovative reporter mouse model that predominantly expresses firefly luciferase (luc2p) in the paw epidermis--the region of murine epidermis that most closely models the tissue architecture of human skin. Combining this animal model with state-of-the-art live animal imaging techniques, we have developed a real-time in vivo analysis work-flow that has allowed us to compare and contrast the efficacies of a wide range nucleic acid-based gene silencing reagents in the skin of live animals. While inhibition was achieved with all of the reagents tested, only the commercially available "self-delivery" modified Accell-siRNAs (Dharmacon) produced potent and sustained in vivo gene silencing. Together, these findings highlight just how informative reliable reporter mouse models can be when assessing novel therapeutics in vivo. Using this work-flow, we developed a novel clinically-relevant topical formulation that facilitates non-invasive epidermal delivery of unmodified and "self-delivery" siRNAs. Remarkably, a sustained >40% luc2p inhibition was observed after two 1-hour treatments with Accell-siRNAs in our topical formulation. Importantly, our ability to successfully deliver siRNA molecules topically brings these novel RNAi-based therapeutics one-step closer to clinical use.

Antibody screening & identification in the general patient population at a tertiary care hospital in New Delhi, India.

BACKGROUND & OBJECTIVES: The development of alloantibodies can significantly complicate transfusion therapy and results in difficulties in cross-matching of blood. Most literature on alloimmunization is limited to multitransfused individuals, with very few studies on the general hospital patients. This study was aimed at assessing the frequency and type of unexpected red cell antibodies in the general patient population at a multispecialty tertiary care centre in New Delhi, India. METHODS: The results of 49,077 antibody screening tests carried out on patients, from January 2009 to December 2012 were analyzed. The clinical and transfusion records were reviewed. The data were compiled and statistically analysed. RESULTS: A total of 49,077 (29,917; 60.96% males and 19,160; 39.04% females) patient samples were screened for the presence of unexpected antibodies. Antibody screening was positive in 403 patients (0.82%). In the serum samples of 164 patients only autoantibodies were identified, 27 revealed autoantibodies with one or more underlying alloantibodies, while 212 patients had only alloantibody/ies in their serum. The overall alloimmunization rate was 0.49 per cent. Antibodies against the Rh system were the most frequent (64.1%), the most common alloantibody identified being anti E (37.2%), followed by anti D (19.2%). INTERPRETATION & CONCLUSIONS: Since clinically significant antibodies are frequently detected in our patient population, antibody screening and if required, identification is the need of the hour. Since antibodies against the common Rh and Kell blood group antigens are the most frequent, provision of Rh and Kell matched red cells may be of protective value.

Siglec-E promotes ß2-integrin-dependent NADPH oxidase activation to suppress neutrophil recruitment to the lung.

Siglec-E is a sialic acid-binding Ig-like lectin expressed on murine myeloid cells. It has recently been shown to function as a negative regulator of ß2-integrin-dependent neutrophil recruitment to the lung following exposure to lipopolysaccharide (LPS). Here, we demonstrate that siglec-E promoted neutrophil production of reactive oxygen species (ROS) following CD11b ß2-integrin ligation with fibrinogen in a sialic acid-dependent manner, but it had no effect on ROS triggered by a variety of other stimulants. Siglec-E promotion of ROS was likely mediated via Akt activation, because siglec-E-deficient neutrophils plated on fibrinogen exhibited reduced phosphorylation of Akt, and the Akt inhibitor, MK2206, blocked fibrinogen-induced ROS. In vivo imaging showed that siglec-E also promoted ROS in acutely inflamed lungs following exposure of mice to LPS. Importantly, siglec-E-promoted ROS were required for its inhibitory function, as the NADPH oxidase inhibitor, apocynin, reversed the siglec-E-mediated suppression of neutrophil recruitment and blocked neutrophil ROS production in vitro. Taken together, these results demonstrate that siglec-E functions as an inhibitory receptor of neutrophils via positive regulation of NADPH oxidase activation and ROS production. Our findings have implications for the inhibitory role of siglec-9 on human neutrophils in sepsis and acute lung injury.

Seroprevalence of anti-HCV antibodies among blood donors of north India.

BACKGROUND & OBJECTIVES: Transfusion of blood and blood products although considered as a life saving treatment modality, but may lead to certain infectious and non-infectious complications in the recipients. The purpose of this analysis was to monitor the seroprevalence of anti-HCV antibody in the blood donor population in a hospital based blood bank in north India, to evaluate the trends over the years (2001-2011). METHODS: Relevant information of all the blood donors who donated whole blood at the department of Transfusion Medicine, Indraprastha Apollo Hospitals, New Delhi from the January 1, 2001 to December 31, 2011 was retrieved from the departmental records. The number of donors who were found reactive for anti-HCV anatibodies was calculated. RESULTS: Of the 2,06,022 blood donors, 1,93,661 were males and 12,361 were females. The percentage of whole blood donors found seroreactive for anti-HCV antibodies was 0.39 per cent (n=795). The seroprevalence of anti-HCV in male blood donors was 0.38 per cent (n=750) and the respective seroprevalence in female blood donors was 0.36 per cent (n=45). No significant change in the trend of HCV seroprevalence was observed over the period under consideration. Maximum seroprevalence of anti-HCV was observed in the age group of 18 to 30 yr (0.41%) and the minimum in the age group of 51 to 60 yr (0.26%). INTERPRETATION & CONCLUSION: HCV seroprevalence in our study was 0.39 per cent and a decreasing trend with age was observed. No significant change in the trend of anti-HCV seroprevalence was seen over a decade. Since, no vaccine is presently available for immunization against HCV infection, transfusion transmitted HCV infection remains a potential threat to the safety of the blood supply.

Antitumor activity and prolonged survival by carbon-nanotube-mediated therapeutic siRNA silencing in a human lung xenograft model.

Carbon nanotubes are novel nanomaterials that are thought to offer potential benefits to a variety of biomedical and clinical applications. In this study, the treatment of a human lung carcinoma model in vivo using siRNA sequences leading to cytotoxicity and cell death is carried out using either cationic liposomes (DOTAP:cholesterol) or amino-functionalized multi-walled carbon nanotubes (MWNT - NH(+)(3)). Validation for the most cytotoxic siRNA sequence using a panel of human carcinoma and murine cells reveals that the proprietary siTOX sequence is human specific and can lead to significant cytotoxic activities delivered both by liposome or MWNT - NH(+)(3) in vitro. A comparative study using both types of vector indicates that only MWNT - NH(+)(3):siRNA complexes administered intratumorally can elicit delayed tumor growth and increased survival of xenograft-bearing animals. siTOX delivery via the cationic MWNT - NH(+)(3) is biologically active in vivo by triggering an apoptotic cascade, leading to extensive necrosis of the human tumor mass. This suggests that carbon-nanotube-mediated delivery of siRNA by intratumoral administration leads to successful and statistically significant suppression of tumor volume, followed by a concomitant prolongation of survival of human lung tumor-bearing animals. The direct comparison between carbon nanotubes and liposomes demonstrates the potential advantages offered by carbon nanotubes for the intracellular delivery of therapeutic agents in vivo. The present work may act as the impetus for further studies to explore the therapeutic capacity of chemically functionalized carbon nanotubes to deliver siRNA directly into the cytoplasm of target cells and achieve effective therapeutic silencing in various disease indications where local delivery is feasible or desirable.