G-protein coupled receptor kinase 5 mediates lipopolysaccharide-induced NFκB activation in primary macrophages and modulates inflammation in vivo in mice.

G-protein coupled receptor kinase-5 (GRK5) is a serine/threonine kinase discovered for its role in the regulation of G-protein coupled receptor signaling. Recent studies have shown that GRK5 is also an important regulator of signaling pathways stimulated by non-GPCRs. This study was undertaken to determine the physiological role of GRK5 in Toll-like receptor-4-induced inflammatory signaling pathways in vivo and in vitro. Using mice genetically deficient in GRK5 (GRK5(-/-) ) we demonstrate here that GRK5 is an important positive regulator of lipopolysaccharide (LPS, a TLR4 agonist)-induced inflammatory cytokine and chemokine production in vivo. Consistent with this role, LPS-induced neutrophil infiltration in the lungs (assessed by myeloperoxidase activity) was markedly attenuated in the GRK5(-/-) mice compared to the GRK5(+/+) mice. Similar to the in vivo studies, primary macrophages from GRK5(-/-) mice showed attenuated cytokine production in response to LPS. Our results also identify TLR4-induced NFκB pathway in macrophages to be selectively regulated by GRK5. LPS-induced IκBα phosphorylation, NFκB p65 nuclear translocation, and NFκB binding were markedly attenuated in GRK5(-/-) macrophages. Together, our findings demonstrate that GRK5 is a positive regulator of TLR4-induced IκBα-NFκB pathway as well as a key modulator of LPS-induced inflammatory response.

Aeroallergens in clinical practice of allergy in India- ARIA Asia Pacific Workshop report.

Allergic diseases such as bronchial asthma, allergic rhinitis and atopic dermatitis are dramatically increasing all over the world including developing countries like India. Today, more than 30% of the population is known to suffer from one or other allergic ailment. Major causative agents implicated are pollen grains, fungal spores, dust mites, insect debris, animal epithelia, etc. Several aerobiological studies have been conducted in different parts of the country to ascertain aerial concentration and seasonality of pollen grains and fungi. Recently, an "All India Coordinated Project on Aeroallergens and Human Health" was undertaken by us to discover the quantitative and qualitative prevalence of aerosols at 18 different centers in the country. Allergenically important airborne pollen identified by clinico-immunologic evaluation are Alnus, Amaranthus, Argemone, Brassica, Cannabis, Cassia, Cedrus, Chenopodium, Cocos, Holoptelia, Mallotus, Morus, Parthenium, Prosopis juliflora, Quercus, Ricinus communis, and grasses such as Cenchrus, Cynodon, Imperata, Pennisetum etc. Cross-reactivity of the IgE antibodies is a common phenomenon among various pollen allergens. Ricinus communis pollen a commonly growing weed/shrub in India, cross-reacts with latex (Hevea brasiliensis), Mercurialis annua and also with seeds of Ricinus communis--all belonging to family Euphorbiaceae but geographically distantly located. Areca catechu cross-reacts with other members of Arecaceae such as Phoenix sylvestris, Cocos nucifera and Borassus flabelifer while pollen of Holoptelia integrifolia from India cross reacts with pollen of Parietaria judaica from Mediterranean Europe, both of which are members of family Urticaceae. Several reports on pollen and fruit syndrome have been analyzed. Experiments conducted by us revealed that pollutants (NO2 and SO2) not only affect pollen morphology but also changes its allergenic potency.

Hypersensitivity to pollen of four different species of Brassica: a clinico-immunologic evaluation in patients of respiratory allergy in India.

BACKGROUND: Rapeseed-mustard is the second most important source of edible oil in India. Several species of Brassica are grown in different parts of country for its oilseeds. OBJECTIVE: The objective was to investigate allergenicity to antigenic extracts of pollen of 4 species of Brassica. METHODS: Brassica campestris, Brassica juncea, Brassica nigra, and Brassica napus were selected for the detailed investigation. Pollen samples from each of the four species were collected from the polliniferous materials. The antigenic and allergenic profiles of these extracts were evaluated by means of sodium dodecyl sulfate-polyacrylamide gel electrophoresis, Skin prick test, enzyme linked immuno sorbent assay and Western blot on atopic individuals. RESULTS: Out of the 159 atopic subjects tested, 21.38% were positive to at least one or other species of Brassica pollen, with highest skin positivity (13.20%) to B. campestris extract. Raised IgE with significant linear correlation with intensity of skin reactions was obtained. Protein fractions of 20, 25, 32, 37, 56, and 90 kDa were recognized by B. campestris and B. juncea whereas 56, 76, 87, and 90 kDa were recognized by B. nigra and B. napus as major IgE binding protein fractions. The patients also showed positivity to other inhalant pollen allergens tested. CONCLUSION: IgE mediated hypersensitivity varied from 4.40% to 13.20% in Indian atopic subjects to pollen of one or the other species of Brassica. Protein fractions of 47, 56, 76, 87, and 90 kDa were identified as IgE binding by all the four species, however individual heterogeneity exists. Thus a local species may be more pertinent for immunotherapy. The major allergen needs to be further characterized.

Soluble and nonsoluble protein assay for antigenic extracts from pollen and seeds of mustard (Brassica spp.).

A study on heterogeneity in water-soluble and non-water-soluble protein profiles of different species of Brassica pollen, seeds, and industrial flour for efficient allergy detection and immunotherapy has been lacking in India. The purpose of this study was to examine heterogeneity in the protein profile of antigenic extracts of different species of pollen, seeds, and seed flour of Brassica. A comparison of water-soluble and non-water-soluble protein profiles of seed extracts was studied. Water-soluble and non-water-soluble proteins were extracted, concentration was estimated by Lowry's method, and biochemical characterization by sodium dodecyl sulfate-polyacrylamide gel electrophoresis was analyzed. Pollen from four species, four species of Brassica seeds comprising 25 varieties, and industrial seed flour were included for water-soluble studies, whereas four species of seeds were included for non-water-soluble protein extraction. Significant variation in protein content was observed among four different species of pollen, seeds, and industrial seed flour, respectively. No significant variation was observed in non-water-soluble extracts of four species or among water-soluble and non-water-soluble content of seeds. Heterogeneity in the protein profile of different species of pollen was not observed. However, variation in banding pattern of water-soluble as well as non-water-soluble protein extracts among four different species of seeds was observed. Interestingly, the industrial seed flour also showed a rich protein banding pattern. Variation in protein content as well as protein profile among different species of seeds of Brassica is recorded.