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Activation of the human innate immune system by Spirulina: augmentation of interferon production and NK cytotoxicity by oral administration of hot water extract of Spirulina platensis.

Spirulina platensis is a cyanobacterial species that is surmised to potentiate the immune system leading to suppression of cancer development and viral infection. Here, we identified the molecular mechanism of the human immune potentiating capacity of Spirulina by analyzing blood cells of volunteers with pre and post oral administration of hot water extract of Spirulina. NK functions represented by IFN gamma production and cytolysis were enhanced after administration of Spirulina in >50% subjects. IFN gamma was produced in an IL-12/IL-18-dependent fashion. In vitro stimulation of blood cells with BCG cell wall skeleton (CWS) allowed more potent IL-12 p40 production in cells from volunteers given Spirulina than in cells without pre-exposure to Spirulina. As BCG-CWS serves as a ligand for Toll-like receptor (TLR) 2 and 4 to raise the maturation stage of monocytes/macrophages, Spirulina may be involved in the signaling responses through Toll in blood cells even when orally administered. These observations indicated that in humans Spirulina acts directly on myeloid lineages and either directly or indirectly on NK cells. The presence of co-operative IL-12 and IL-18 is critically important for NK-mediated IFN gamma production.

Administration, Oral↗

Revealing the complementation of ferredoxin by cytochrome b (5) in the Spirulina- (6)-desaturation reaction by N-terminal fusion and co-expression of the fungal-cytochrome b (5) domain and Spirulina- (6)-acyl-lipid desaturase.

Spirulina-acyl-lipid desaturases are integral membrane proteins found in thylakoid and plasma membranes. These enzymes catalyze the fatty acid desaturation process of Spirulina to yield gamma-linolenic acid (GLA) as the final desaturation product. It has been reported that the cyanobacterial desaturases use ferredoxin as an electron donor, whereas the acyl-lipid desaturase in plant cytoplasm and the acyl-CoA desaturase of animals and fungi use cytochrome b (5). The low level of ferredoxin present in Escherichia coli cells leads to an inability to synthesize GLA when the cells are transformed with the Spirulina-(6) desaturase, desD, and grown in the presence of the reaction substrate, linoleic acid. In this study, Spirulina-(6) desaturase, encoded by the desD gene, was N-terminally fused and co-expressed with the cytochrome b (5) domain from Mucor rouxii. The product, GLA, made heterologously in E. coli and Saccharomyces cerevisiae, was then detected and analyzed. The results revealed the production of GLA by Spirulina-(6) desaturase fused or co-expressed with cytochrome b (5) in E. coli cells, in which GLA production by this gene cannot occur in the absence of cytochrome b (5). Moreover, the GLA production ability in the E. coli host cells was lost after the single substitution mutation was introduced to H52 in the HPGG motif of the cytochrome b (5) domain. These results revealed the complementation of the ferredoxin requirement by the fusion or co-expression of the fungal-cytochrome b (5) domain in the desaturation process of Spirulina-(6) desaturase. Furthermore, the free form of cytochrome b (5) domain can also enhance GLA production by the Spirulina-desD gene in yeast cells.

Amino Acid Motifs↗

The expression of three desaturase genes of Spirulina platensis in Escherichia coli DH5alpha. Heterologous expression of Spirulina-desaturase genes.

The genes from a cyanobacterium--Spirulina platensis strain C1--that encode the acyl-lipid desaturases (desC, desA and desD) involved in gamma-linolenic (GLA) synthesis have been successfully expressed for the first time in Escherichia coli by employing a pTrcHisA expression system. In this report, the authors describe the expression of the three Spirulina N-terminal 6xHis-desaturases as well as the functional analysis of these recombinant proteins. The gene products of desC, desA and desD have approximate molecular masses of 37, 45, and 47 kDa, respectively. Enzymatic activity measurement of these products was carried out in vivo to demonstrate that (i) the expressed proteins are in functional form, and (ii) the cofactors of the host system can complement the system of Spirulina platensis. The study demonstrated that the gene products of desC and desA catalyzed the reactions in vivo where the enzyme substrates were provided in appropriate concentration. This indicates that the delta9 and delta12 desaturases were expressed in the heterologous host in their active form, and that these two reactions can be carried out in an E. coli host cell using its cofactors system. In contrast, delta6 desaturase activity can be detected only in vitro where electron carriers are provided. This suggests that while this enzyme is expressed in the heterologous host in its active form, its function in vivo is suppressed, as the electron carriers of the host system cannot complement the system of Spirulina platensis.

Bacterial Proteins↗

[Role of sodium ions and their uptake by cells of cultured blue-green algae, Spirulina platensis and Spirulina maxima].

The growth of the blue-green algae Spirulina platensis and Spirulina maxima, cultured in complete mineral Zarouk medium containing Na+ or Na+-deficient medium, was studied over a period of 24 h. The optical densities of S. platensis and S. maxima cells, determined during the last hour of exposure to sodium deficiency, amounted to 55.6 and 32.6%, respectively, of the optical densities of the same cells grown in complete Zarouk medium. Moreover, the cultures grown in Na+-deficient medium exhibited increased ability to take up sodium (which was low in S. platensis and S. maxima cells cultured in complete mineral medium). It is concluded that the two species studied are characterized by periodic, on the order of minutes, changes in the cellular uptake and release of sodium.

Biological Transport↗

[Nutritive value of the spirulina algae (Spirulina maxima)].

Nine experiments were conducted, five of them in vivo to determine the limiting amino acids and digestibility of spiruline algae for the rat, and four in vitro to determine the digestibility of the product in pepsin and ruminal liquid. None of the amino acids studied (lysine, methionine, histidine) added alone or in combination to 10% protein (either crude or true) diets provided exclusively by spiruline, seems to be limiting although the results could be masked by the low palatability and acceptability of the product by the rats. The apparent digestibility of the algae was 67.4%. For the in vitro tests, the algae were subjected to several physical or chemical treatments, and the digestibility of the resulting product determined by four different techniques. In no case did the tested treatments have any effect on its digestibility.

Amino Acids, Essential↗

Efficacy of spirulina extract plus zinc in patients of chronic arsenic poisoning: a randomized placebo-controlled study.

BACKGROUND: Millions of people in Bangladesh, India, Taiwan, and Chile are consuming high concentration of arsenic through drinking water, and thousands of them have already developed chronic arsenic poisoning. There is no specific treatment. Some authors suggest the use of vitamins and minerals for more than 6 months. The present placebo-controlled double-blind study was conducted to evaluate effectiveness of spirulina extract plus zinc in the treatment of chronic arsenic poisoning. METHODS: Forty-one patients of chronic arsenic poisoning were randomly treated orally by either placebo (17 patients) or spirulina extract (250 mg) plus zinc (2 mg) (24 patients) twice daily for 16 weeks. Each patient was supplied with arsenic-safe drinking water by installing a locally made water filter at household level. Effectiveness of spirulina extract plus zinc was evaluated by comparing changes in skin manifestations (clinical scores), arsenic contents in urine and hair, between the placebo- and spirulina extract plus zinc-treated groups. RESULTS: The concentrations of total arsenic in water (without filtration) of placebo- and spirulina extract plus zinc-treated groups were 150.1 +/- 18.3 and 161.7 +/- 23.9 microg/l, respectively. Intake of these high concentrations of arsenic lead to increased excretion of arsenic in urine (72.1 +/- 14.5 microg/l in placebo-treated group and 78.4 +/- 19.1 microg/l in spirulina plus zinc-treated group). After 2 weeks of using filtered water, there were significant reduction of both arsenic intake through water and urinary arsenic excretion (8.3 +/- 3.6 microg/l and 18.4 +/- 7.3 microg/l in placebo group; 9.7 +/- 5.4 microg/l and 21.6 +/- 5.8 microg/l) in spirulina extract plus zinc-treated group. There was a sharp increase in urinary excretion of arsenic (138 +/- 43.6 microg/l) at 4 weeks following spirulina plus zinc administration and the effect was continued for another 2 weeks. Spirulina extract plus zinc removed 47.1% arsenic from scalp hair. Spirulina extract had no major adverse effect that required physician's attention. The clinical scores (median) for melanosis before and after treatment with placebo was not statistically significant (p > 0.05), whereas in spirulina extract plus zinc-treated group it was statistically significant (p < 0.01). In cases of keratosis, the median clinical scores before and after treatment was not statistically significant (p > 0.05) in placebo-treated group. In spirulina extract plus zinc-treated group, the clinical scores for keratosis before and after treatment was statistically significant (p < 0.05). CONCLUSIONS: Results show that spirulina extract (250 mg) plus zinc (2 mg) twice daily for 16 weeks may be useful for the treatment of chronic arsenic poisoning with melanosis and keratosis.

Adolescent↗

The nutritional value of dehydrated, blue-green algae (Spirulina platensis) for poultry.

Three experiments were conducted to evaluate the nutritional value of the blue-green algae, Spirulina platensis, grown on a synthetic media. In Experiment 1, day-old, White Leghorn cockerel chicks (120) were fed isonitrogenous diets containing 0, 5, 10, 15, and 20% of dried spirulina. At 3 wk of age, the growth of the chicks fed 10 and 20% of spirulina was depressed (P less than .05), although feed efficiency was not affected. In Experiment 2, 250 1-day-old, Hubbard by Hubbard, male broiler chicks were fed experimental diets containing 0, 1.5, 3.0, 6.0, or 12.0% of spirulina for 41 days. Although the growth of the chicks fed the spirulina diets was not different from that of the chicks receiving the control diet, the birds receiving the 12% spirulina diet grew slower (P less than .05) than the chicks fed all of the other spirulina diets. In Experiment 3, 600 1-wk-old, Japanese qual were used to study the effects of 0, 1.5, 3.0, 6.0, and 12.0% of spirulina on growth, egg production, egg quality, fertility, hatchability, and the growth of the F1 generation of dams fed Spirulina. There were no significant differences due to the spirulina content in any of the parameters studied-except for yolk color, which increased with each succeeding level of spirulina, and for fertility, which was higher for all spirulina treatments versus the control.

Animal Feed↗

[Incorporation of a spiruline (Spirulina maxima) in swine food].

Two experiments on early weaned piglets and one experiment on sows during the first two reproductive cycles were made to determine the nutritive value of Spirulina maxima algae in the pig, taken as a "nutritional model" for man. The algae were introduced into the diets of the piglets, between 12 and 42 days of age, in a proportion representing 12 p. 100 of the total proteins. This level corresponded to 12 p. 100 algae in the first age feed (up to 21 days) in replacement of skim milk, and 8 p. 100 of the second age feed in replacement of soyabean meal. For the overall experimental period, from 12 to 42 days, the apparent digestibilities of the diets, measured by the chronium oxide marking, decreased by 6.9 and 6.1 points, respectively, for energy and total crude proteins as compared to the control diet. Replacement of the same raw materials by a soluble fish protein concentrate only led to a reduction of 2.9 and 3.5 points for energy and proteins. However, during the first week following weaning, the fish concentrate and spirulina gave identical results which were lower than those of the control (fig. 1). In spite of this lowering in the apparent digestibility the growths of the piglets were satisfactory and equivalent for all groups. Thus, the mĕtabolic utilisation of the fraction of feed absorbed was better for the spirulina group than for the control group, notably during the period between 12 to 21 days, although the supply of lysine in the spirulina group was 12 p. 100 lower (tab. VII). Consequently, this partially compensates for the low apparent digestibility of the algae in the young animal. For the female, the spirulina algaes were continously administered in a proportion of 5 p. 100 of the diet from a live weight of 75 kg. This did not change either the age or the weight at onset of puberty (tab IX). During the first reproductive cycle, the net weight gain of the sows receiving spirulina was lower than that of the control sows, but the number of piglets and the weight of the litter were higher (tab X). During the second cycle, performances were almost identical for the two groups, but piglets from the spirulina sows were slightly heavier than those from the control sows, in corelation with smaller litters (tab XI). During lactation, the lipid content of the milk was not substancially modified by addition of spirulina to the diet (tab. XIII). At the end of the two reproductive cycle the characteristics of the two groups of sows were identical, with a slightly higher culling rate for the spirulina group, but without direct relationship with the use of algae. Furthermore, the productivity of the sows was the same: 16.34 piglets weaned per sow and per year for the controls and 15.99 for the spirulina sows. Thus, if considering spirulina as a protein supplement to traditional proteins, its use in pig feeding does not seem to give rise to particular problem. However, it is advisable to restrict their supply to a level not exceeding 25 p...

Age Factors↗

Spirulina attenuates cyclosporine-induced nephrotoxicity in rats.

Cyclosporine (CsA) causes a dose-related decrease in renal function in experimental animals and humans. The generation of reactive oxygen species (ROS) has been implicated in CsA-induced nephrotoxicity. It was previously shown that Spirulina, a blue-green algae, with antioxidant properties effectively attenuated the doxorubicin-induced cardiotoxicity in mice and cisplatin-induced nephrotoxicity in rat. The present study investigated the nephroprotective role of Spirulina against CsA-induced nephrotoxicity in rats. Spirulina (500 mg kg(-1) b.w.) was administered orally for 3 days before and 14 days concurrently with CsA (50 mg kg-1 b.w.). Rats treated with CsA showed nephrotoxicity as evidenced from a significant elevation in plasma urea, creatinine, urinary N-acetyl-beta-D-glucosaminidase (beta-NAG) and a decrease in creatinine and lithium clearance. Pretreatment with Spirulina protected the rats from CsA-induced nephrotoxicity. The CsA-induced rise in plasma urea and creatinine and the decrease in creatinine and lithium clearance were attenuated by Spirulina. There was a significant increase in plasma and kidney tissue MDA with CsA. Spirulina prevented the rise in plasma and kidney tissue MDA. Histopathology of the kidney from CsA-treated rats showed severe isometric vacuolization and widening of the interstitium. However, pretreatment with Spirulina prevented such changes, and the kidney morphology was comparable to that of the control. Spirulina treatment did not alter the blood CsA levels. These results suggest that Spirulina has a protective effect against nephrotoxicity induced by CsA. This study further supports the crucial role of the antioxidant nature of Spirulina in protecting against CsA-induced oxidative stress.

Acetylglucosaminidase↗

Studies on the preventive effect of Spirulina maxima on fatty liver development induced by carbon tetrachloride, in the rat.

The aim of the present work was to assess if the feeding of either the oil extract of Spirulina maxima or of its defatted fraction would prevent fatty liver development, induced in rats by a single intraperitoneal dose of carbon tetrachloride (CCl4). Liver and serum lipids were evaluated 4 days after treatment with this agent. Concentration of liver lipids did not differ in rats fed on a purified diet either without or with one of the fractions of Spirulina, except for total cholesterol, which showed a slight increase in the group receiving the oil extract of Spirulina. However, after CCl4 treatment, liver total lipids and triacylglycerols were significantly lower in rats fed on a diet containing any fraction of Spirulina (defatted or the oil fraction) than in rats without Spirulina in their diet. Furthermore, the increased liver cholesterol values, induced by CCl4 treatment, were not observed in rats receiving Spirulina. In addition, rats receiving whole Spirulina in their diet and treated only with the vehicle showed an increase in the percentage of HDL values. The changes in VLDL and LDL induced by CCl4 treatment were not observed in the whole Spirulina group. Furthermore, after CCl4 treatment the values of the liver microsomal thiobarbituric acid-reactive substances were lower in the whole Spirulina group than in the control group. These results support the potential hepatoprotective role of Spirulina.

Analysis of Variance↗

Credentials of Spirulina diet on stability and flux related properties on the biomineralization process during oxalate mediated renal calcification in rats.

BACKGROUND: High Spirulina diet is a potential risk factor for nephrolithiasis since it has the capacity to increase urinary oxalate and uric acid level, facilitating lithogenesis. Our aim was to identify the effect of Spirulina diet during hyperoxaluric condition in Wistar albino rats. METHODS: The animals were divided into four groups: control (Gl, n=6); ethylene glycol (EG) induced (G2, n=6); EG+Spirulina (G3, n=6); Spirulina alone (G4, n=6). EG at 0.75% was administered to G2 and G3 through drinking water for 4 weeks and Spirulina 1500 mg/kg feed was administered to G3 and G4. RESULTS: Urinary parameters like oxalate, uric acid, calcium, urea, and creatinine (P<0.001) were found increased after Spirulina diet under hyperoxaluric conditions compared to the same without Spirulina diet. Similarly the BUN, plasma contents of uric acid, urea, creatinine (P<0.001) were found to be raised in G3. The renal and RBC GSH levels, as estimated by HPLC, seemed decreased when compared to G2. CONCLUSIONS: The present study shows that free radicals aid in the progression of nephrolithiasis. The crystal deposition was found to be high in the renal cells of G3 than G2 and TEM revealed damage in renal cell of G3 implying that the disease deteriorates by free radical injury. In contrast the Spirulina diet alone (G4) did not induce any features relating to stone forming condition suggesting that free radical release might have been suppressed due to enrichment of dietary antioxidants and vitamins. Thus the present investigation demonstrates that during hyperoxaluric conditions the Spirulina diet must possibly be avoided and can be considered in normal subjects checked for family history of renal stone deposition.

Animals↗

Inhibitory effect of mast cell-mediated immediate-type allergic reactions in rats by spirulina.

We investigated the effect of spirulina on mast cell-mediated immediate-type allergic reactions. Spirulina dose-dependently inhibited the systemic allergic reaction induced by compound 48/80 in rats. Spirulina inhibited compound 48/80-induced allergic reaction 100% with doses of 100-1000 microg/g body weight, i.p. Spirulina (10-1000 microg/g body weight, i.p.) also significantly inhibited local allergic reaction activated by anti-dinitrophenyl (DNP) IgE. When rats were pretreated with spirulina at a concentration ranging from 0.01 to 1000 microg/g body weight, i.p., the serum histamine levels were reduced in a dose-dependent manner. Spirulina (0.001 to 10 microg/mL) dose-dependently inhibited histamine release from rat peritoneal mast cells (RPMC) activated by compound 48/80 or anti-DNP IgE. The level of cyclic AMP in RPMC, when spirulina (10 microg/mL) was added, transiently and significantly increased about 70-fold at 10 sec compared with that of control cells. Moreover, spirulina (10 microg/mL) had a significant inhibitory effect on anti-DNP IgE-induced tumor necrosis factor-alpha production. These results indicate that spirulina inhibits mast cell-mediated immediate-type allergic reactions in vivo and in vitro.

Animals↗

Antioxidant and antiproliferative activities of Spirulina and Chlorella water extracts.

Liver fibrosis is a chronic liver disease that will further develop to cirrhosis if severe damage continues to form. A potential treatment for liver fibrosis is to inhibit activated hepatic stellate cell (HSC) proliferation and, subsequently, to induce HSC apoptosis. It has been reported that antioxidants are able to inhibit the proliferation of HSCs. In this study, the aqueous extract of spirulina was chosen as the source of antioxidant to investigate the inhibitory effect on the proliferation of HSC. The growth inhibitory effects of aqueous spirulina and chlorella extract on human liver cancer cells, HepG2, were also studied and compared in pairs. Results indicated that the total phenol content of spirulina was almost five times greater than that of chlorella (6.86 +/- 0.58 vs 1.44 +/- 0.04 mg tannic acid equivalent/g of algae powder, respectively). The antioxidant activity of spirulina determined by the ABTS*+ method was higher than chlorella (EC50: 72.44 +/- 0.24 micromol of trolox equivalent/g of spirulina extract vs 56.09 +/- 1.99 micromol of trolox equivalent/g of chlorella extract). Results of DPPH* assay also showed a similar trend as the ABTS*+ assay (EC50: 19.39 +/- 0.65 micromol of ascorbic acid equivalent/g of spirulina extract vs 14.04 +/- 1.06 micromol of ascorbic acid equivalent/g of chlorella extract). The aqueous extracts of these two algae both showed antiproliferative effects on HSC and HepG2, but spirulina was a stronger inhibitor than chlorella. Annexin-V staining showed that aqueous extract of spirulina induced apoptosis of HSC after 12 h of treatment. In addition, the aqueous extract of spirulina triggered a cell cycle arrest of HSC at the G2/M phase.

Animals↗

Effects of a Spirulina-based dietary supplement on cytokine production from allergic rhinitis patients.

Spirulina represents a blue-green alga that is widely produced and commercialized as a dietary supplement for modulating immune functions, as well as ameliorating a variety of diseases. We have previously shown that the in vitro culture of Spirulina with human peripheral blood mononuclear cells (PBMCs) modulated the production of cytokines. In the present study, we evaluated the impact of a Spirulina-based dietary supplement (Earthrise Nutritionals, Inc., Irvine, CA) on patients with allergic rhinitis by assessing the production of cytokines [interleukin (IL)-4, interferon (IFN)-gamma, and IL-2] critical in regulating immunoglobulin E-mediated allergy. In a randomized double-blinded crossover study versus placebo, allergic individuals were fed daily with either placebo or Spirulina, at 1,000 mg or 2,000 mg, for 12 weeks. PBMCs isolated before and after the Spirulina feeding were stimulated with phytohemagglutinin (PHA) prior to determining the levels of cytokine from cell culture supernatants. Although Spirulina seemed to be ineffective at modulating the secretion of Th1 cytokines (IFN-gamma and IL-2), we discovered that Spirulina, administered at 2,000 mg/day, significantly reduced IL-4 levels by 32% from PHA-stimulated cells. These results indicate that Spirulina can modulate the Th profile in patients with allergic rhinitis by suppressing the differentiation of Th2 cells mediated, in part, by inhibiting the production of IL-4. To our knowledge, this is the first human feeding study that demonstrates the protective effects of Spirulina towards allergic rhinitis.

Adolescent↗