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In vivo effects of traditional Ayurvedic formulations in Drosophila melanogaster model relate with therapeutic applications.

Dwivedi V, Anandan EM, Mony RS, Muraleedharan TS, Valiathan MS, Mutsuddi M, Lakhotia SC - PLoS ONE (2012)

Bottom Line: On the contrary, feeding larvae on normal food and adults on AR supplement had no effect on fecundity but a comparable regime of feeding on RS-supplemented food improved fecundity.RS feeding did not cause heavy metal toxicity.Thus, Drosophila, with its very rich genetic tools and well-worked-out developmental pathways promises to be a very good model for examining the cellular and molecular bases of the effects of different Ayurvedic formulations.

View Article: PubMed Central - PubMed

Affiliation: Cytogenetics Laboratory, Department of Zoology, Banaras Hindu University, Varanasi, India.

ABSTRACT

Background: Ayurveda represents the traditional medicine system of India. Since mechanistic details of therapy in terms of current biology are not available in Ayurvedic literature, modern scientific studies are necessary to understand its major concepts and procedures. It is necessary to examine effects of the whole Ayurvedic formulations rather than their "active" components as is done in most current studies.

Methods: We tested two different categories of formulations, a Rasayana (Amalaki Rasayana or AR, an herbal derivative) and a Bhasma (Rasa-Sindoor or RS, an organo-metallic derivative of mercury), for effects on longevity, development, fecundity, stress-tolerance, and heterogeneous nuclear ribonucleoprotein (hnRNP) levels of Drosophila melanogaster using at least 200 larvae or flies for each assay.

Results: A 0.5% (weight/volume) supplement of AR or RS affected life-history and other physiological traits in distinct ways. While the size of salivary glands, hnRNP levels in larval tissues, and thermotolerance of larvae/adult flies improved significantly following feeding either of the two formulations, the median life span and starvation resistance improved only with AR. Feeding on AR or RS supplemented food improved fecundity differently. Feeding of larvae and adults with AR increased the fecundity while the same with RS had opposite effect. On the contrary, feeding larvae on normal food and adults on AR supplement had no effect on fecundity but a comparable regime of feeding on RS-supplemented food improved fecundity. RS feeding did not cause heavy metal toxicity.

Conclusions: The present study with two Ayurvedic formulations reveals formulation-specific effects on several parameters of the fly's life, which seem to generally agree with their recommended human usages in Ayurvedic practices. Thus, Drosophila, with its very rich genetic tools and well-worked-out developmental pathways promises to be a very good model for examining the cellular and molecular bases of the effects of different Ayurvedic formulations.

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Larval salivary glands attain greater size following feeding on 0.5% AR (B) or RS (C) than in normally fed larvae (A) of same age (spiracle eversion stage).Individual polytene nuclei are also bigger in larvae reared on AR- (E) or RS-supplemented (F) food than in those fed on regular food (D); nuclei marked with arrows in D–F are shown at higher magnification in insets (also see Table 2). Histograms in G show the mean (±S.E.) DAPI fluorescence (in arbitrary fluorescence units on Y-axis) in distal polytene nuclei in salivary glands from larvae reared on regular (CON) or formulation (AR or RS) supplemented food (numbers of nuclei examined in each case are indicated in parentheses above the respective bars. Scale bar in B applies to A–C while that in D, applies to D–F.
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pone-0037113-g003: Larval salivary glands attain greater size following feeding on 0.5% AR (B) or RS (C) than in normally fed larvae (A) of same age (spiracle eversion stage).Individual polytene nuclei are also bigger in larvae reared on AR- (E) or RS-supplemented (F) food than in those fed on regular food (D); nuclei marked with arrows in D–F are shown at higher magnification in insets (also see Table 2). Histograms in G show the mean (±S.E.) DAPI fluorescence (in arbitrary fluorescence units on Y-axis) in distal polytene nuclei in salivary glands from larvae reared on regular (CON) or formulation (AR or RS) supplemented food (numbers of nuclei examined in each case are indicated in parentheses above the respective bars. Scale bar in B applies to A–C while that in D, applies to D–F.

Mentions: In order to see if the AR or RS-supplement in food affected internal larval organs, we examined the different internal organs of late 3rd instar larvae. Various organs like the gut, imaginal discs, brain ganglia etc in larvae reared on the formulation supplemented food appeared generally comparable in size and disposition to those in larvae fed on regular food. Their SG, however, appeared somewhat larger in the formulation fed larvae. Therefore, we compared the dimensions (width and length) of SG of late third instar (spiracle eversion stage) larvae that were fed on regular (Fig. 3A) or 0.5% AR (Fig. 3B) or 0.5% RS (Fig. 3C) supplemented food since hatching. It is interesting that the length and width of the SG are significantly enhanced (Table 3 and Fig. 3A–C) in larvae grown on formulation supplemented food. The number of cells in each of the larval salivary glands is fixed during embryonic stage and the subsequent growth of this tissue occurs through endo-replication cycles that are highly regulated in relation to the anatomical location of each nucleus in the gland [16], [17], [18]. Therefore, we wanted to know if the increased dimensions of these glands correlated with change in nuclear size and increased DNA content. In order to obtain SG of same age, larvae that had just everted their anterior spiracles were selected and immediately dissected in Poels' salt solution (PSS) [19]. Measurement of diameters of nuclei in the posterior most 5–7 cells in these glands revealed that the nuclear size is significantly greater in the AR- (Fig. 3E) or RS-fed (Fig. 3F) larval SG compared to that in normally fed control larvae (Fig. 3D). Measurement of DNA-specific DAPI fluorescence of distal nuclei in salivary glands from larvae that had just begun the spiracle eversion (Fig. 3D–F, Table 3) revealed that increased nuclear size is paralleled by increase in the DNA content in these polytene nuclei. These effects are more pronounced in AR-fed larvae than in those receiving the RS supplement (Table 3 and Fig. 2G). These observations show that while the larvae reared on formulation supplemented food take a few hours less to pupate than those reared on regular food (see Fig. 2 above), the polytene nuclei in their SG undergo greater numbers of endoreplication cycles so that in parallel with increased DNA content per nucleus, the nuclear and SG dimensions also increase.


In vivo effects of traditional Ayurvedic formulations in Drosophila melanogaster model relate with therapeutic applications.

Dwivedi V, Anandan EM, Mony RS, Muraleedharan TS, Valiathan MS, Mutsuddi M, Lakhotia SC - PLoS ONE (2012)

Larval salivary glands attain greater size following feeding on 0.5% AR (B) or RS (C) than in normally fed larvae (A) of same age (spiracle eversion stage).Individual polytene nuclei are also bigger in larvae reared on AR- (E) or RS-supplemented (F) food than in those fed on regular food (D); nuclei marked with arrows in D–F are shown at higher magnification in insets (also see Table 2). Histograms in G show the mean (±S.E.) DAPI fluorescence (in arbitrary fluorescence units on Y-axis) in distal polytene nuclei in salivary glands from larvae reared on regular (CON) or formulation (AR or RS) supplemented food (numbers of nuclei examined in each case are indicated in parentheses above the respective bars. Scale bar in B applies to A–C while that in D, applies to D–F.
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC3351451&req=5

pone-0037113-g003: Larval salivary glands attain greater size following feeding on 0.5% AR (B) or RS (C) than in normally fed larvae (A) of same age (spiracle eversion stage).Individual polytene nuclei are also bigger in larvae reared on AR- (E) or RS-supplemented (F) food than in those fed on regular food (D); nuclei marked with arrows in D–F are shown at higher magnification in insets (also see Table 2). Histograms in G show the mean (±S.E.) DAPI fluorescence (in arbitrary fluorescence units on Y-axis) in distal polytene nuclei in salivary glands from larvae reared on regular (CON) or formulation (AR or RS) supplemented food (numbers of nuclei examined in each case are indicated in parentheses above the respective bars. Scale bar in B applies to A–C while that in D, applies to D–F.
Mentions: In order to see if the AR or RS-supplement in food affected internal larval organs, we examined the different internal organs of late 3rd instar larvae. Various organs like the gut, imaginal discs, brain ganglia etc in larvae reared on the formulation supplemented food appeared generally comparable in size and disposition to those in larvae fed on regular food. Their SG, however, appeared somewhat larger in the formulation fed larvae. Therefore, we compared the dimensions (width and length) of SG of late third instar (spiracle eversion stage) larvae that were fed on regular (Fig. 3A) or 0.5% AR (Fig. 3B) or 0.5% RS (Fig. 3C) supplemented food since hatching. It is interesting that the length and width of the SG are significantly enhanced (Table 3 and Fig. 3A–C) in larvae grown on formulation supplemented food. The number of cells in each of the larval salivary glands is fixed during embryonic stage and the subsequent growth of this tissue occurs through endo-replication cycles that are highly regulated in relation to the anatomical location of each nucleus in the gland [16], [17], [18]. Therefore, we wanted to know if the increased dimensions of these glands correlated with change in nuclear size and increased DNA content. In order to obtain SG of same age, larvae that had just everted their anterior spiracles were selected and immediately dissected in Poels' salt solution (PSS) [19]. Measurement of diameters of nuclei in the posterior most 5–7 cells in these glands revealed that the nuclear size is significantly greater in the AR- (Fig. 3E) or RS-fed (Fig. 3F) larval SG compared to that in normally fed control larvae (Fig. 3D). Measurement of DNA-specific DAPI fluorescence of distal nuclei in salivary glands from larvae that had just begun the spiracle eversion (Fig. 3D–F, Table 3) revealed that increased nuclear size is paralleled by increase in the DNA content in these polytene nuclei. These effects are more pronounced in AR-fed larvae than in those receiving the RS supplement (Table 3 and Fig. 2G). These observations show that while the larvae reared on formulation supplemented food take a few hours less to pupate than those reared on regular food (see Fig. 2 above), the polytene nuclei in their SG undergo greater numbers of endoreplication cycles so that in parallel with increased DNA content per nucleus, the nuclear and SG dimensions also increase.

Bottom Line: On the contrary, feeding larvae on normal food and adults on AR supplement had no effect on fecundity but a comparable regime of feeding on RS-supplemented food improved fecundity.RS feeding did not cause heavy metal toxicity.Thus, Drosophila, with its very rich genetic tools and well-worked-out developmental pathways promises to be a very good model for examining the cellular and molecular bases of the effects of different Ayurvedic formulations.

View Article: PubMed Central - PubMed

Affiliation: Cytogenetics Laboratory, Department of Zoology, Banaras Hindu University, Varanasi, India.

ABSTRACT

Background: Ayurveda represents the traditional medicine system of India. Since mechanistic details of therapy in terms of current biology are not available in Ayurvedic literature, modern scientific studies are necessary to understand its major concepts and procedures. It is necessary to examine effects of the whole Ayurvedic formulations rather than their "active" components as is done in most current studies.

Methods: We tested two different categories of formulations, a Rasayana (Amalaki Rasayana or AR, an herbal derivative) and a Bhasma (Rasa-Sindoor or RS, an organo-metallic derivative of mercury), for effects on longevity, development, fecundity, stress-tolerance, and heterogeneous nuclear ribonucleoprotein (hnRNP) levels of Drosophila melanogaster using at least 200 larvae or flies for each assay.

Results: A 0.5% (weight/volume) supplement of AR or RS affected life-history and other physiological traits in distinct ways. While the size of salivary glands, hnRNP levels in larval tissues, and thermotolerance of larvae/adult flies improved significantly following feeding either of the two formulations, the median life span and starvation resistance improved only with AR. Feeding on AR or RS supplemented food improved fecundity differently. Feeding of larvae and adults with AR increased the fecundity while the same with RS had opposite effect. On the contrary, feeding larvae on normal food and adults on AR supplement had no effect on fecundity but a comparable regime of feeding on RS-supplemented food improved fecundity. RS feeding did not cause heavy metal toxicity.

Conclusions: The present study with two Ayurvedic formulations reveals formulation-specific effects on several parameters of the fly's life, which seem to generally agree with their recommended human usages in Ayurvedic practices. Thus, Drosophila, with its very rich genetic tools and well-worked-out developmental pathways promises to be a very good model for examining the cellular and molecular bases of the effects of different Ayurvedic formulations.

Show MeSH
Related in: MedlinePlus