Savneet Kaur

Distinct alleles of mannose-binding lectin (MBL) and surfactant proteins A (SP-A) in patients with chronic cavitary pulmonary aspergillosis and allergic bronchopulmonary aspergillosis.

Abstract Background: Distinct host immune status predisposes to different forms of pulmonary aspergillosis. Methods: Patients with chronic cavitary pulmonary aspergillosis (CCPA; n=15) or allergic bronchopulmonary aspergillosis (ABPA; n=7) of Caucasian origin were screened for single nucleotide polymorphisms (SNPs) in the collagen region of surfactant proteins A1 (SP-A1) and A2 (SP-A2) and mannose binding lectin (MBL). Results: The T allele at T1492C and G allele at G1649C of SP-A2 were observed at slightly higher frequencies in ABPA patients (86% and 93%) than in controls (63% and 83%), and the C alleles at position 1492 and 1649 were found in higher frequencies in CCPA patients (33% and 25%) than in ABPA patients (14% and 7%) (all p>0.05). However, the CC genotype at position 1649 of SP-A2 was significantly associated with CCPA (χ2=7.94; pcorr≤0.05). Similarly, ABPA patients showed a higher frequency of the TT genotype (71%) at 1492 of SP-A2 than controls (43%) and CCPA patients (41%) (p>0.05). In the case of MBL, the T allele (OR=3.1, range 1.2–8.9; p≤0.02) and CT genotype (χ2=6.54; pcorr≤0.05) at position 868 (codon 52) were significantly associated with CCPA, but not with ABPA. Further analysis of genotype combinations at position 1649 of SP-A2 and at 868 of MBL between patient groups showed that both CC/CC and CC/CT SP-A2/MBL were found only in CCPA patients, while GG/CT SP-A2/MBL was significantly higher in CCPA patients in comparison to ABPA patients (p≤0.05). SNPs analysed in SP-A1 did not differ between cases and controls. Conclusions: Distinct alleles, genotypes and genotype combinations of SP-A2 and MBL may contribute to differential susceptibility of the host to CCPA or ABPA. Clin Chem Lab Med 2007;45:183–6.

Protective role of mannan‐binding lectin in a murine model of invasive pulmonary aspergillosis

Innate immune molecules such as lung collectins and serum pentraxins have evolved as important host defence proteins against Aspergillus fumigatus, a medically important opportunistic fungal pathogen. Mannan‐binding lectin (MBL), an opsonin and lectin complement pathway activator, constitutes another vital player of innate immunity against several pathogenic organisms in the serum. Studies have reported significant binding of MBL to A. fumigatus; however, the protective role of MBL against A. fumigatus‐mediated invasive disease remains elusive. Henceforth, we investigated the contribution of externally administered recombinant human (rh) MBL towards anti‐fungal defence in invasive pulmonary aspergillosis (IPA) by in vivo and in vitro studies. In murine models of IPA with corticosteroid‐induced immunosuppression, rhMBL‐treated mice showed 80% survival compared to untreated IPA mice with no survivors. Treated IPA mice also showed a marked increase in tumour necrosis factor (TNF)‐α and interleukin (IL)‐1α and a significant decrease in pulmonary fungal hyphae and IL‐10. In vitro, rhMBL‐bound A. fumigatus conidia showed a dose‐dependent increase in the deposition of C4b, the first product of the lectin pathway. There was an enhanced uptake of A. fumigatus conidia by the polymorphonuclear cells (PMNs) in the presence of rhMBL that increased further in the presence of MBL supplemented with MBL‐deficient serum. However, an increase in the oxidative burst of PMNs and A. fumigatus killing were observed only when MBL was supplemented with MBL‐deficient serum. The study suggests a therapeutic role of ex vivo‐administered MBL in host defence against aspergillosis, possibly through MBL‐mediated complement activation and other protective mechanisms aimed both directly at the pathogen, and indirectly through modulation of the host inflammatory responses.

Role of collectins in innate immunity against aspergillosis

The protective role of lung surfactant proteins SP-A, SP-D and MBL in the host defense against both allergic and invasive aspergillosis was identified and established by a series of in vitro and in vivo studies. Therapeutic administration of SP-D and MBL proteins in a murine model of pulmonary invasive aspergillosis rescued mice from death. In mice mimicking human allergic bronchopulmonary aspergillosis, SP-A and SP-D suppressed IgE levels, eosinophilia, pulmonary cellular infiltration and cause a marked shift from a pathogenic Th2 to a protective Th1 cytokine profile. SP-A and SP-D knock-out mice studies made significant contributions in understanding the mechanisms by which SP-A and SP-D modulate the host defense response in patients suffering from pulmonary allergies and infections. The results suggested that individuals with any structural or functional defects in these innate immune molecules due to genetic variations might be susceptible to aspergillosis. SNPs in SP-A2 and MBL genes showed significant associations with patients of allergic bronchopulmonary aspergillosis in an Indian population. The patients carrying either one or both of GCT and AGG alleles of SP-A2 and patients with A allele at position 1011 of MBL had markedly higher eosinophilia, total IgE antibodies and lower FEV1 (the clinical markers of ABPA). Our results show that collectins play an important role in Aspergillus mediated allergies and infections.

Biofilm formation by Aspergillus fumigatus

Aspergillus fumigatus is a well adapted, opportunistic fungus that causes a severe and commonly fatal disease, invasive pulmonary aspergillosis (IPA), in highly immunocompromised patients, aspergilloma in patients with lung cavities and allergic bronchopulmonary aspergillosis (ABPA) in hypersensitive individuals. Recent studies have suggested that biofilm formation by A. fumigatus may be one of the most important virulence factors in IPA and aspergilloma. Several fungal constituents may contribute to the formation of biofilm structures on host cells, including cell wall components, secondary metabolites and drug transporters. The biofilm phenotype of the fungus is refractory to most conventional antifungal treatment options. Thus, an in-depth analysis and understanding of A. fumigatus biofilms is necessary to devise newer and better antifungal targets for treating complex A. fumigatus biofilm-associated diseases.

Hepatic Progenitor Cells in Action: Liver Regeneration or Fibrosis?

Liver injury caused by drugs, viruses, and toxins that impede the proliferation of mature hepatocytes results in the activation of hepatic progenitor cells (HPCs), which then participate in the restoration of the damaged liver tissue. HPCs are known to be bipotential cells, capable of forming both hepatocytes and cholangiocytes when regeneration by mature hepatocytes is plagued or impaired. Both clinical studies of liver disease and certain experimental animal models of liver injury conspicuously show the presence of activated HPC response and proliferation. However, in addition to regeneration, the proliferation of HPCs also determines the appearance of a ductular reaction that has been correlated with progressive portal fibrosis, suggesting intricate links between activation of HPCs and fibrogenesis. The current review highlights the role of activated HPCs in both hepatic regeneration and fibrosis during liver injury.

When nano meets stem : The impact of nanotechnology in stem cell biology

Nanotechnology and biomedical treatments using stem cells are among the latest conduits of biotechnological research. Even more recently, scientists have begun finding ways to mate these two specialties of science. The advent of nanotechnology has paved the way for an explicit understanding of stem cell therapy in vivo and by recapitulation of such in vivo environments in the culture, this technology seems to accommodate a great potential in providing new vistas to stem cell research. Nanotechnology carries in its wake, the development of highly stable, efficient and specific gene delivery systems for both in vitro and in vivo genetic engineering of stem cells, use of nanoscale systems (such as microarrays) for investigation of gene expression in stem cells, creation of dynamic three-dimensional nano-environments for in vitro and in vivo maintenance and differentiation of stem cells and development of extremely sensitive in vivo detection systems to gain insights into the mechanisms of stem cell differentiation and apoptosis in different disease models. The present review presents an overview of the current applications and future prospects for the use of nanotechnology in stem cell biology.

Angiogenesis in liver regeneration and fibrosis: “a double-edged sword”

Angiogenesis, defined as the formation of new microvasculature from preexisting blood vessels and mature endothelial cells, plays a major role in wound healing and scar formation, and it is associated with inflammatory responses. Angiogenesis can occur in physiological conditions, such as during liver regeneration, and in pathological situations, such as during the progression of fibrosis to cirrhosis and also during tumor angiogenesis. Cellular cross-talk among liver sinusoidal endothelial cells (LSECs), hepatic stellate cells and hepatocytes is believed to play an important role in the angiogenesis process during both liver regeneration and development of cirrhosis. In addition to mature endothelial cells, bone marrow (BM)-derived circulating endothelial progenitor cells (EPCs) have been recently identified for their contribution to post-natal vasculogenesis/angiogenesis. In vivo, EPCs are mobilized into the peripheral blood in response to tissue ischemia or traumatic injury, migrate to the sites of injured endothelium and differentiate into mature endothelial cells. In our recent studies, we have explored the role of EPC-mediated angiogenesis in liver regeneration and/or cirrhosis. Results have demonstrated significantly increased endogenous levels of circulating EPCs in cirrhotic patients in comparison to the controls. Also, EPCs from cirrhotic patients have been observed to stimulate substantial angiogenesis by resident LSECs in vitro via paracrine factors such as vascular endothelial growth factor and platelet-derived growth factor. This review gives an overview of the angiogenesis process in liver regeneration and disease and discusses a new mechanism for intrahepatic angiogenesis mediated by BM-derived EPCs.

Transfection of Endothelial Nitric Oxide Synthase Gene Improves Angiogenic Efficacy of Endothelial Progenitor Cells in Rabbits with Hindlimb Ischemia

Background: The present study explored the effect of endothelial nitric oxide synthase (eNOS) gene transfer on the angiogenic potential of ex vivo expanded endothelial progenitor cells (EPCs) in a rabbit model of hindlimb ischemia. Methods: Rabbit peripheral blood EPCs were cultured and transfected with mammalian expression vector pcDNA3.1-eNOS containing full-length human eNOS gene. Ischemia was induced in the right hind limb of three groups of rabbits by ligation of the distal external iliac artery and excision of the common and superficial femoral arteries. In one group of animals, ten days after the surgery, autologous eNOS-EPCs were transplanted intramuscularly in the ischemic limb. Two other groups received an equivalent number of unmodified EPCs or phosphate buffered saline (PBS) respectively. Results: Two weeks after cell transplantation, the in vivo expression of eNOS was detected in limb tissue sections of eNOS-EPCs treated animals. Animals treated with eNOS-EPCs had a significant reduction in ischemic muscle necrosis and inflammation, augmentation in the capillary density (P< 0.05) and angiographic scores demonstrating distal arterial reconstitution and enhanced angiogenesis in comparison to animals transplanted with EPCs or PBS (P< 0.05). Conclusion: We conclude that modification of EPCs by eNOS constitutes an effective strategy to improve the efficacy of EPCs for therapeutic angiogenesis.

Association of SP-D, MBL and I-NOS genetic variants with pulmonary tuberculosis

Background : Pulmonary tuberculosis is caused by Mycobacterium tuberculosis . It is a multifactorial disease with both host as well as pathogen factors contributing to susceptibility and protection from the disease. Various reports have highlighted important roles of lung surfactant protein D (SP-D), mannan-binding lectin (MBL) and I-NOS in innate immune defense against M. tuberculosis Aims : The present study investigated the role of polymorphisms in three candidate genes encoding Lung surfactant protein D, Mannan binding lectin and Inducible Nitric oxide synthase, in susceptibility and protection to pulmonary tuberculosis. Settings and Design : A case-control association study of SNPs in lung surfactant protein D (SP-D), mannan-binding lectin (MBL) and I-NOS with pulmonary tuberculosis in Indian population was carried out. This involved sequencing of all the coding exons of lung surfactant protein D (SP-D) , while, exon 1 (collagen region) and exon 4 (carbohydrate recognition domain) of mannan-binding lectin (MBL) and exons 2, 8 and 16 of I-NOS and their flanking intronic regions for single nucleotide polymorphisms in DNA samples isolated from 30 pulmonary tuberculosis patients and 30 controls of Indian population. Statistical analysis: Various allele frequencies were calculated using online two by two table (home.clara.net/sisa/). Odds ratio and P values were calculated at 95% confidence interval (CI). Results : A total of fourteen single nucleotide polymorphisms (5 in SP-D , 5 in MBL and 4 in I-NOS ) were observed of which four (G459A SP-D , G274T I-NOS , G1011A and T357G MBL ) have not been reported earlier. Four single nucleotide polymorphisms viz. G459A of exon 7 of SP-D ( P =0.00, odds ratio (OR) = 4.96, 2.18 P = 0.00 or= 3.85 1.66 P =0.00 or=4.04, 2.20 P =0.00 or=4.46, 2.40 Conclusion: The present study has led to identification of 4 SNPs in SP-D , MBL and I-NOS associated with pulmonary tuberculosis in Indian population.

To study the effect of curcumin on the growth properties of circulating endothelial progenitor cells

Curcumin has been shown to regulate the expression of genes implicated in tumor cell proliferation, metastasis, chemotherapy resistance, and angiogenesis. Endothelial progenitor cells (EPCs) have been recently described in the peripheral blood as cells contributing to both physiological and pathological angiogenesis. In the current study, we evaluated the effect of curcumin on these angiogenic cells. EPCs were isolated, expanded, and characterized ex vivo. These cells were then treated with different concentrations of curcumin. The formation of EPC colonies in culture and their proliferation was analyzed by 5′-bromo-2′-deoxyuridine assays in absence and presence of curcumin. Further, the expression of two important cell cycle inhibitory proteins, p21 and p53, in the curcumin- and culture medium-treated cells without curcumin was evaluated by intracellular flow cytometry. The results showed that there was a significant decrease in the formation of EPC colonies in culture. EPC proliferation was significantly inhibited by curcumin in a dose-dependent manner. Flow cytometry analysis showed a twofold increase in the expression of both p21 and p53 in curcumin-treated cells as compared to the medium-treated cells, suggesting that curcumin inhibits EPC growth by mainly inhibiting the G1 to S phase transition in the cell cycle. It would be further worthwhile to study the effect of curcumin on EPC-mediated angiogenic activity.