Ratan Kumar

Safety and healing efficacy of Sea buckthorn (Hippophae rhamnoides L.) seed oil on burn wounds in rats

The present investigation was undertaken to determine the safety and efficacy of supercritical CO2-extracted Hippophae rhamnoides L. (Sea buckthorn) (SBT) seed oil on burn wound model. SBT seed oil was co-administered by two routes at a dose of 2.5 ml/kg body weight (p.o.) and 200 microl (topical) for 7 days on experimental burn wounds in rats. The SBT seed oil augmented the wound healing process as indicated by significant increase in wound contraction, hydroxyproline, hexosamine, DNA and total protein contents in comparison to control and reference control treated with silver sulfadiazine (SS) ointment. Histopathological findings further confirmed the healing potential of SBT seed oil. SBT seed oil treatment up-regulated the expression of matrix metalloproteinases (MMP-2 and 9), collagen type-III and VEGF in granulation tissue. It was observed that SBT seed oil also possesses antioxidant properties as evidenced by significant increase in reduced glutathione (GSH) level and reduced production of reactive oxygen species (ROS) in wound granulation tissue. In acute and sub-acute oral toxicity studies, no adverse effects were observed in any of the groups administered with SBT seed oil. These results suggest that the supercritical CO2-extracted Sea buckthorn seed oil possesses significant wound healing activity and have no associated toxicity or side effects.

Cytoprotective and antioxidant activity of Rhodiola imbricata against tert-butyl hydroperoxide induced oxidative injury in U-937 human macrophages

The present study reports cytoprotective and antioxidant activity of aqueous and alcoholic extracts of Rhodiolaimbricata rhizome on tert-butyl hydroperoxide (tert-BHP) induced cytotoxicity in U-937 human macrophages. There was an increase in cytotoxicity and apoptosis significantly in the presence of tert-BHP over control cells. The tert-BHP induced cytotoxicity can be attributed to enhanced reactive oxygen species (ROS) production which in turn is responsible for fall in reduced glutathione (GSH) levels; further there was a significant decrease in mitochondrial potential and increase in apoptosis and DNA fragmentation. Both aqueous and alcoholic extracts of Rhodiola rhizome at a concentration of 250 μg/ml were found to inhibit tert-BHP induced free radical production, apoptosis and to restore the anti-oxidant levels to that of the control cells. The alcoholic extract of Rhodiola showed higher cytoprotective activities than aqueous extract. These observations suggest that the alcoholic and aqueous extracts of Rhodiola have marked cytoprotective and antioxidant activities.

A poly-herbal formulation accelerates normal and impaired diabetic wound healing.

In the present study, a poly‐herbal formulation (PHF) was prepared by combining the aqueous lyophilized leaf extracts of Hippophae rhamnoides L. and Aloe vera L. and the ethanol rhizome extract of Curcuma longa L., in an optimized ratio (1 : 7 : 1). The efficacy of PHF treatment was studied in normal and impaired diabetic rats using a full‐thickness cutaneous wound model. Topical PHF treatment increased cellular proliferation and collagen synthesis at the wound site in normal rats, as evidenced by the significant increase in DNA, total protein, hydroxyproline, and hexosamine contents in comparison with a positive control treated with a povidone–iodine ointment. The histological examinations and matrix metalloproteinases expression also correlated well with the biochemical findings, confirming the efficacy of PHF in normal wounds. In the streptozotocin‐induced diabetic rats, PHF treatment increased hydroxyproline and hexosamine content. A faster wound contraction was also observed in PHF‐treated normal and diabetic rats. The PHF also promoted angiogenesis as evidenced by an in vitro chick chorioallantoic membrane model and in vivo up‐regulated vascular endothelial growth factor expression. The results suggest that PHF possesses significant wound healing potential in both normal as well as chronic diabetic wounds.

A preclinical study of the effects of seabuckthorn (Hippophae rhamnoides L.) leaf extract on cutaneous wound healing in albino rats.

Hippophae rhamnoides L. (family Elaeagnaceae), commonly known as seabuckthorn, is a wild shrub growing at high altitude (1200-4500 meters) in adverse climatic conditions. The aim of the present study was to evaluate healing potential of seabuckthorn leaves in a preclinical study on rats using a cutaneous excision-punch wound model. Four full-thickness excision-type wounds of 8.0 mm diameter were created on the dorsal surface of rats under aseptic conditions. The aqueous lyophilized extract of seabuckthorn leaves, at doses of 0.5%, 1.0%, and 1.5% w/v prepared in propylene glycol, were applied topically twice daily for 7 days. Control animals received the vehicle alone in an identical manner. Wound granulation tissues were excised on eighth day postwounding, and the hydroxyproline, hexosamine, total protein content, and antioxidant levels were determined. Wound surface area was also measured on the eighth day before wound excision to determine wound contraction. Topical application of 1.0% seabuckthorn leaf extract statistically significantly augmented the healing process, as evidenced by increases in the content of hydroxyproline and protein as well as the reduction in wound area when compared with similar effects in response to treatment using povidone-iodine ointment (standard care). The reduced glutathione, vitamin C, superoxide dismutase, catalase, and glutathione peroxidase activities showed significant increases in seabuckthorn leaf extract-treated wounds as compared to controls. The lipid peroxide levels were significantly decreased in leaf extract-treated wounds. The results suggest that aqueous leaf extract of seabuckthorn promotes wound healing, which may be due to increased antioxidant levels in the granulation tissue.

Angiotensin converting enzyme insertion allele in relation to high altitude adaptation

Angiotensin converting enzyme (ACE) gene I/D polymorphism has been associated with high altitude (HA) disorders as well as physical performance. We, however, envisage that the polymorphism may be associated with adaptation to the hypobaric hypoxia of altitude, thus facilitating physical performance. For this purpose, three unrelated adult male groups, namely (1) the Ladakhis (HLs), who reside at and above a height of 3600 m, (2) lowlanders, who migrated to Ladakh (MLLs), and (3) resident lowlanders (LLs), have been investigated. The HLs had significantly (p < 0.001) greater numbers of the II homozygotes and the ID heterozygotes than the DD homozygotes, the genotype distribution being 0.46, 0.43 and 0.11 for II, ID and DD genotypes respectively. The MLLs comprised 60% II homozygotes, which was higher (p < 0.001) than the HLs (46%). In the LLs, the heterozygotes were greater (p < 0.001) in number than the II and DD homozygotes. The I allele frequency was 0.72 in the MLLs, 0.67 in the HLs and 0.55 in the LLs. Polymorphism study suggested that the II genotype could be associated with altitude adaptation, which might influence physical efficiency.

Mechanism of Wound-Healing Activity of Hippophae rhamnoides L. Leaf Extract in Experimental Burns

The present investigation was undertaken to evaluate the healing efficacy of lyophilized aqueous leaf extract of Sea buckthorn (Hippophae rhamnoides L., family Elaeagnaceae) (SBT) and to explore its possible mechanism of action on experimental burn wounds in rats. The SBT extract, at various concentrations, was applied topically, twice daily for 7 days. Treatment with silver sulfadiazine (SSD) ointment was used as reference control. The most effective concentration of the extract was found to be 5.0% (w/w) for burn wound healing and this was further used for detailed study. The SBT-treated group showed faster reduction in wound area in comparison with control and SSD-treated groups. The topical application of SBT increased collagen synthesis and stabilization at the wound site, as evidenced by increase in hydroxyproline, hexosamine levels and up-regulated expression of collagen type-III. The histological examinations and matrix metalloproteinases (MMP-2 and -9) expression also confirmed the healing efficacy of SBT leaf extract. Furthermore, there was significant increase in levels of endogenous enzymatic and non-enzymatic antioxidants and decrease in lipid peroxide levels in SBT-treated burn wound granulation tissue. The SBT also promoted angiogenesis as evidenced by an in vitro chick chorioallantoic membrane model and in vivo up-regulated vascular endothelial growth factor (VEGF) expression. The SBT leaf extract had no cytotoxic effect on BHK-21 cell line. In conclusion, SBT aqueous leaf extract possesses significant healing potential in burn wounds and has a positive influence on the different phases of wound repair.

Influence of sea buckthorn (Hippophae rhamnoides L.) flavone on dermal wound healing in rats

The present investigation was undertaken to determine the efficacy of topical administration of flavone of sea buckthorn (Hippophae rhamnoides L.) on cutaneous wound healing in rats. Four full-thickness excision wounds were created on the back of rat and 1.0% w/v flavone prepared in propylene glycol was applied topically. Control animals received the vehicle alone in an identical manner. The healing of the wound was assessed by the rate of wound contraction, period of epithelialization, hydroxyproline, hexosamine, antioxidants estimation and histopathology of the granulation tissue. The sea buckthorn flavone promoted the wound healing activity as indicated by improved rate of wound contraction, decreased time taken for epithelialization (16.3 days versus 24.8 days in controls) and significant increase in hydroxyproline (26.0%) and hexosamine (30.0%) content. These findings were also confirmed by histopathological examinations. In addition, it was observed that sea buckthorn flavone possesses potent antioxidant properties as evidenced by significant increase in reduced glutathione (55.0%), vitamin C (70.0%) and catalase (20.0%) activities in wound granulation tissue. The flavone treatment also resulted in significant decrease in lipid peroxide levels (39.0%). The results suggest that the sea buckthorn flavone promotes wound healing activity.

Modulatory effects of seabuckthorn (Hippophae rhamnoides L.) in hypobaric hypoxia induced cerebral vascular injury.

Cerebral edema caused by vascular leakage is a major problem in various injuries of the CNS, such as stroke, head injury and high-altitude illness. A common feature of all these disorders is the fact that they are associated with tissue hypoxia. Hypoxia has been suggested to be a major pathogenic factor for the induction of vascular leakage in the brain. The objective of the present study was to evaluate potential of seabuckthorn (SBT) (Hippophae rhamnoides L.) seed oil in curtailing hypoxia induced transvascular fluid leakage in brain of hypoxia-exposed rats. Exposure of animals to hypobaric hypoxia (9144 m, 5h) caused a significant increase in the transvascular leakage studied by measuring water content and leakage of sodium fluorescein dye in the brain. Hypoxic stress also significantly enhanced the oxidative stress markers such as free radicals and malondialdehyde and it accompanied with decreased levels of antioxidants such as glutathione, glutathione peroxidase and superoxide dismutase. Pretreatment of animals with SBT seed oil significantly restricted the hypoxia induced increase in fluorescein dye leakage suggesting protection against hypoxia induced transvascular leakage in the brain. Hypoxia induced increase in the levels of free radicals and malondialdehyde were significantly lowered after SBT pretreatment. The SBT seed oil pretreatment also resulted in the significantly improved hypoxic tolerance as evidenced by increased hypoxic gasping time and survival time and decreased plasma catecholamine levels, as compared to hypoxic animals. These observations suggest that SBT seed oil possesses significant hypoxia protection activity and curtailed hypoxia induced enhanced vascular leakage in the brain.

A dose dependent adaptogenic and safety evaluation of Rhodiola imbricata Edgew, a high altitude rhizome

To examine the dose dependent adaptogenic activity aqueous extract of Rhodiola imbricata root was orally administered in rats at different doses, 30 min prior to cold (5 degrees C)-hypoxia (428 mm Hg)-restraint (C-H-R) exposure. The maximal effective adaptogenic dose of the extract was 100 mg/kg body weight. The acute and sub-acute toxicity of the extract was also studied in rats. Sub-acute toxicity studies included administration of single oral dose of 1 g/kg and 2 g/kg of extract once daily for 14 days and maximal effective single oral dose of 100 mg/kg once daily for 30 days. At the end of each treatment period the biochemical parameters related to liver function, kidney function, lipids (triglycerides, cholesterol) and hematological parameters were estimated in serum and blood. Biochemical and hematological analysis showed no significant changes in any of the parameters examined in treated groups animal, in comparison to control animals. No significant change was observed in organ weight/body weight ratios, of any organ studied in comparison to control rats. The oral LD(50) of the extract was observed to be >10 g/kg, indicating an adequate margin of safety. No histopathological changes were observed in the vital organs studied of the treated animals. These results suggest that aqueous extract of R. imbricata root possess potent adaptogenic activity with no acute and sub-acute toxicity.

Anti-oxidative effect of Rhodiola imbricata root extract in rats during cold, hypoxia and restraint (C-H-R) exposure and post-stress recovery.

Anti-oxidative potential of Rhodiola imbricata root aqueous extract was examined in rats, administered orally at a dose of 100 mg/kg both in single and multiple doses, 30 min prior to cold (5 degrees C)-hypoxia (428 mmHg)-restraint (C-H-R) exposure. Lipid per-oxidation, anti-oxidant parameters and lactate dehydrogenase (LDH), were studied in blood, liver and muscle of rats on attaining T(rec)23 degrees C during C-H-R exposure and after recovery (T(rec)37 degrees C) from C-H-R induced hypothermia. The results of untreated control rats on attaining T(rec)23 degrees C showed a significant increase in blood, liver and muscle malondialdehyde (MDA) and LDH levels. Hepatic catalase (CAT) and muscle glutathione S-transferase (GST) also increased significantly. Administration of single dose of Rhodiola imbricata root aqueous extract significantly restricted rise in blood MDA, increased blood reduced glutathione (GSH) and superoxide dismutase (SOD) activity with restricted rise in blood, liver and muscle LDH; improved liver and muscle SOD on attaining T(rec)23 degrees C and T(rec)37 degrees C; liver CAT on attaining T(rec)23 degrees C and liver GST during recovery. Multiple doses treatment of the extract further increased blood, liver and muscle GSH and GST levels; restricted increase in LDH on attaining T(rec)23 degrees C and recovery; increased CAT during recovery. Results suggested the anti-oxidant potential of Rhodiola root extract during C-H-R exposure and post-stress recovery and it also maintained cell membrane permeability.