The effects of C-phycocyanin (C-pc), a phycobiliprotein, on the expression of pro-fibrotic mediators in hyper-tropic scarring such as connective tissue growth factor (CTGF) and α-smooth muscle actins (α-SMA) were investigated in relation to trans-differentiation of fibroblast to myo-fibroblast, an icon of scar formation. C-pc was isolated from Spirulina Platensis extract using sonication method and C-pc concentration was determined by Bennet and Bogorad equation. α-SMA and CTGF levels in wounded primary human dermal fibroblasts were determined by western blot analysis and immuno-fluorescence confocal microscope was employed. Fibroblast contractility was examined by three-dimensional collagen lattice contraction assay. There was an elevation of α-SMA (121%) and CTGF (143%) levels in wound cells as compared with non-wound cells. The does-response profiles of down regulation demonstrated that the maximum inhibitions of α-SMA by 63% (p<0.05) and CTGF by 50% (p<0.1) were achieved by C-pc (6 nM) treated cells. In confocal assay, non-wound fibroblasts exhibited basal level of α-SMA staining, while wounded cells without C-pc treatment showed strong up-regulation of α-SMA by 147% (p<0.05). C-pc (6 nM) inhibited α-SMA expression by 70% (p<0.05) and reduced collagen contraction by 29% (p<0.05). C-pc seemed to lessen the over expression of CTGF, α-SMA, subsequently alleviating the fibrotic contracture. This study suggests the potential application of C-pc to regulation of the expression of pro-fibrotic mediators in scarring process and its potential usage as an efficient means for anti-fibrosis therapy.
Actins* ; Blotting, Western ; Cicatrix ; Collagen ; Connective Tissue Growth Factor* ; Connective Tissue* ; Contracture ; Down-Regulation ; Fibroblasts ; Humans ; Methods ; Myofibroblasts ; Phycocyanin* ; Sonication ; Spirulina ; Up-Regulation ; Wound Healing ; Wounds and Injuries

OBJECTIVETo investigate the therapeutic effect of C-phycocyanin (C-PC) from Spirulina platensis on paraquat (PQ)-induced pulmonary fibrosis in rats.

METHODSA total of 90 healthy Wistar rats were randomly and equally divided into normal control group, model group (PQ group), and C-PC treatment group (C-PC group). Each rat in the PQ group and C-PC group were orally administered with a single dose of PQ (50 mg/kg) to establish a rat model of PQ poisoning. Then, the rats in the normal control group and PQ group were orally given saline solution (1 ml/100 g) every day, and the rats in the C-PC group were orally given C-PC (50 mg/kg) every day. Six rats were randomly selected from each group on days 1, 3, 7, 14, and 28. The inferior lobe of each rat's right lung was homogenized for the measurement of hydroxyproline (HYP) and maleic dialdehyde (MDA) levels and superoxide dismutase (SOD) activity. Parts of each rat's left lung were subject to HE staining and Masson staining for pathological observation, and the expression of transforming growth factor-β(1) (TGF-β(1)), nuclear factor-kappa B p65 (NF-κB p65), and tumor necrosis factor-α (TNF-α) in lung tissue was measured by immunohistochemistry.

RESULTSThe HYP levels on days 1, 3, 7, 14, and 28 and MDA levels on days 14 and 28 were significantly lower in the C-PC group than in the PQ group (P < 0.05, P < 0.01). The SOD activity was significantly higher in the C-PC group than in the PQ group on days 1, 7, 14, and 28 (P < 0.05, P < 0.01). The protein content of TGF-β(1) and the activities of NF-κB p65 and TNF-α in the PQ group and C-PC group were significantly higher than those in the normal control group, while the indices in the C-PC group were significantly lower than those in the PQ group (P < 0.05, P < 0.01). The pathological observation showed that C-PC could alleviate pulmonary alveolitis and fibrosis in rats with PQ poisoning.

CONCLUSIONC-PC can significantly inhibit PQ-induced pulmonary alveolitis and fibrosis in rats.

Animals ; Lung ; metabolism ; pathology ; Male ; NF-kappa B ; metabolism ; Paraquat ; poisoning ; Phycocyanin ; pharmacology ; Pulmonary Fibrosis ; chemically induced ; metabolism ; prevention & control ; Rats ; Rats, Wistar ; Transcription Factor RelA ; metabolism ; Transforming Growth Factor beta ; metabolism ; Tumor Necrosis Factor-alpha ; metabolism

We developed large-scale preparation of phycoerythrin from Porphyra haitanensis, a main economic red algae in China. Firstly, P. haitanensis thallus was broken by using "swelling and smash" method. Then times of grads ammonium sulfate precipitation applied to the crude extraction were compared. Desalted solution was further purified with one-step chromatography using hydroxyapatite and properties on spectrum and molecular weight were identified finally. The results indicated that after four times of ammonium sulfate precipitation (15%, 50%, 10% and 40%), the absorption spectrum purity of P. haitanensis achieved 0.9 (A564/A280), and 507.82 mg phycoerythrin (A564/A280 > 3.2) was obtained from 7 kg fresh algae after further hydroxyapatite chromatography. This research provides a potential way for preparation of phycoerythrin in large sclae.
Ammonium Sulfate ; chemistry ; Chromatography ; methods ; Phycoerythrin ; isolation & purification ; Porphyra ; chemistry

PURPOSE: It has been reported that Spirulina, a blue-green algae with potent antioxidant properties, affords significant protection against inflammation and fibrosis in the liver in vivo. The aim of the present study was to establish the possible protective role of C-phycocyanin, one of the active ingredients of Spirulina, in an experimental model of fibrosis in the kidney. METHODS: The study was carried out using male C57BL6 mice. Mice were divided into the following four groups: sham-operated group; C-phycocyanin (PC)-treated sham group; unilateral ureteral obstruction (UUO) group; and PC with UUO group. We evaluated renal TGF-beta mRNA, MCP-1, and osteopontin using real-time RT PCR. We evaluated renal TGF-beta, alpha-SMA, and CD68 by immunohistochemistry. We recorded light microscopic findings of kidney specimens. RESULTS: PC significantly decreased the expression of MCP-1 and alpha-SMA mRNA. Renal gene levels of expression of TGF-beta, MCP-1, and osteopontin in the UUO group were significantly higher than the sham-operated group (p<0.01). The levels of expression of TGF-beta, MCP-1, and osteopontin mRNA of kidneys in the PC-treated UUO group were significantly lower than the untreated UUO group (p< 0.05). The magnitude of expression of TGF-beta and alpha-SMA protein in the kidneys of the PC-treated UUO group was significantly less than the untreated UUO control group (p<0.05). CONCLUSION: The results of the present study suggest that PC has anti-inflammatory and anti-fibrotic effects in an experimental UUO murine model.
Animals ; Cyanobacteria ; Fibrosis ; Humans ; Immunohistochemistry ; Inflammation ; Kidney ; Light ; Liver ; Male ; Mice ; Models, Theoretical ; Osteopontin ; Phycocyanin ; Polymerase Chain Reaction ; RNA, Messenger ; Salicylamides ; Spirulina ; Transforming Growth Factor beta ; Ureteral Obstruction

No abstract available.
Fibrosis ; Inflammation ; Phycocyanin

BACKGROUND AND OBJECTIVES: The thioredoxin (TRx) system is a ubiquitous thiol oxidoreductase pathway that regulates cellular reduction/oxidation status. Although endothelial cell (EC) hypoxic damage is one of the important pathophysiologic mechanisms of ischemic heart disease, its relationship to the temporal expression pattern of the TRx system has not yet been elucidated well. The work presented here was performed to define the expression pattern of the TRx system and its correlation with cellular apoptosis in EC lines in hypoxic stress. These results should provide basic clues for applying aspects of the TRx system as a therapeutic molecule in cardiovascular diseases. SUBJECTS AND METHODS: Hypoxia was induced with 1% O2, generated in a BBL GasPak Pouch (Becton Dickinson, Franklin Lakes, NJ, USA) in human endothelial progenitor cells (hEPC) and human umbilical vein endothelial cells (HUVEC). Apoptosis of these cells was confirmed by Annexin-V: Phycoerythrin flow cytometry. Expression patterns of TRx; TRx reductase; TRx interacting protein; and survival signals, such as Bcl-2 and Bax, in ECs under hypoxia were checked. RESULTS: Apoptosis was evident after hypoxia in the two cell types. Higher TRx expression was observed at 12 hours after hypoxia in hEPCs and 12, 36, 72 hours of hypoxia in HUVECs. The expression patterns of the TRx system components showed correlation with EC apoptosis and cell survival markers. CONCLUSION: Hypoxia induced significant apoptosis and its related active changes of the TRx system were evident in human EC lines. If the cellular impact of TRx expression pattern in various cardiovascular tissues under hypoxia or oxidative stress was studied meticulously, the TRx system could be applied as a new therapeutic target in cardiovascular diseases, such as ischemic heart disease or atherosclerosis.
Anoxia ; Apoptosis ; Atherosclerosis ; Cardiovascular Diseases ; Cell Hypoxia ; Cell Survival ; Endothelial Cells ; Flow Cytometry ; Human Umbilical Vein Endothelial Cells ; Humans ; Lakes ; Myocardial Ischemia ; Oxidative Stress ; Phycoerythrin ; Stem Cells ; Thioredoxins

We studied the effect of photodynamic therapy with phycobiliproteins on human liver cancer cells in vitro. With 3-(4, 5-Dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide assay (MTT assay), we used two phycobiliproteins, R-phycoerythrin (R-PE) and C-phycocyanin (C-PC) prepared from Porphyra yezoensis, to determine the killing rates and apoptosis rates of human liver cancer cells (SMMC-7721) mediated by laser. When the concentration of R-PE was 120 mg/L, the survival rate of human liver cancer cells was 27% after treated by Argon laser with 100 J/cm2 doses, while the survival rate in the control group (without adding R-PE) was 65%. When the C-PC concentration was 120 mg/L, the survival cell rate was 47% after treated by He-Ne laser with 35 J/cm2 dose, while the survival rate in the control group (without adding C-PC) was 70%. After handled only with these two kinds of phycobiliproteins for 72 h, the growth of cancer cells presented significant inhibition. The maximal inhibition rates reached up to 31% with R-PE (120 mg/L concentration) and 27% with C-PC (250 mg/L concentration) respectively. After irradiated by laser for 8 h, the maximal cell apoptosis rates were 31.54% with R-PE and 32.54% with C-PC, respectively. It indicated that R-PE and C-PC extracted from Porphyra yezoensis could develop to new photosensitizers for cancer photodynamic therapy.
Apoptosis ; drug effects ; radiation effects ; Cell Line, Tumor ; Humans ; Lasers ; Liver Neoplasms ; pathology ; Photochemical Processes ; Photochemotherapy ; methods ; Phycobiliproteins ; isolation & purification ; pharmacology ; Phycoerythrin ; isolation & purification ; pharmacology ; Porphyra ; chemistry

Phycocyanin subunits liposomes (PCS-lip) were prepared and its cellular uptake and photodynamic therapy (PDT) effect on cancer cells were studied. In the experiment, film dispersion method was used to prepare phycocyanin subunits liposomes; particle size and distribution were detected by zetasizer and transmission electric microscope; the effects of liposome as carrier on cell uptake in vitro were evaluated in S180 by using fluorescence microscope; and photodynamic therapy effect was assessed with MTT method. As shown in the results, the particle size mainly ranged from 80 nm to 160 nm, and average encapsulation rate was 42.3%. In the concentration of 100 microg x mL(-1), transfection rate reached (18.5 +/- 0.8)% at 2 h, (23.1 +/- 0.9)% at 4 h, keeping a balance in 5-6 h, and its photodynamic therapy effect in vitro improved with the increasing of concentration of phycocyanin subunits liposomes. In the concentration of 200 micro x mL(-1) cell survival rate of BGC-823 and S180 reached (45 +/- 5.2)% and (36 +/- 5.5)%, respectively, and the cell survival rate differentiation between PCS-PDT group and PCS-lip-PDT group reached 7%-11% (P < 0.05). In this study film dispersion method could keep the biological activity of phycocyanin subunits very well. Phycocyanin subunits liposomes will transfect cells more quickly than phycocyanin subunits in the same concentration, and in the same conditions, phycocyanin subunits liposomes have the better PDT effect on cancer cells as they were incubated with cells for 4 h.
Animals ; Cell Line, Tumor ; Cell Survival ; drug effects ; Drug Carriers ; Drug Stability ; Humans ; Liposomes ; chemistry ; Mice ; Particle Size ; Photochemotherapy ; methods ; Phycocyanin ; administration & dosage ; pharmacology ; Protein Subunits ; administration & dosage ; pharmacology ; Sarcoma 180 ; pathology ; Stomach Neoplasms ; pathology

BACKGROUND: Chronic neuropathic pain is often associated with altered immune function and the modulated immune cell response play a role in neuropathic pain by experimental nerve injury. In order to assess the possible changes in lymphocytes function following peripheral mononeuropathy, this study examined the lymphocyte subpopulation of the spleen using the monoclonal antibodies against the membrane surface markers in neuropathic BALB/c mice by a partial transection of sciatic nerve (PST). METHODS: After confirming tactile allodynia by paw withdrawal threshold, the splenic lymphocytes were stained with fluorescein isothiocyanate (FITC)-conjugated anti-mouse CD45R/B220 (B cell) and CD4 (helper/inducer T cell) or with phycoerythrin (PE)-conjugated anti-mouse CD90.2 (total T cell) and CD8 (suppressor/cytotoxic T cell). The proportions of subsets were analyzed using a FACScan laser flow cytometry system on postoperative day 5 and day 18 respectively. RESULTS: PST induced a mechanical allodynia as verified by the von Frey test at both 5 and 8 days postoperatively compared to pre-surgery (P < 0.05). Lymphocyte subpopulation was affected by PST. The proportion of CD4+ subset was significantly larger in the PST group than in the sham operated group on day 5, while the proportion of CD8+ subset was larger on day 18. In the PST group, there were significantchanges in the proportion of CD4+ on day 5 and in the proportion of CD8+ on day 18 (P < 0.05) compared to pre-surgery. There were no significant fluctuations in the proportion of total splenic T cell and B cell subsets of PST group compared to sham operated group. CONCLUSIONS: These results suggest that development of mononeuropathy is responsible for the proportional changes in splenic lymphocyte subsets in mice.
Animals ; Antibodies, Monoclonal ; B-Lymphocyte Subsets ; Flow Cytometry ; Fluorescein ; Hyperalgesia ; Lymphocyte Subsets* ; Lymphocytes* ; Membranes ; Mice* ; Mononeuropathies ; Neuralgia ; Peripheral Nerve Injuries* ; Peripheral Nerves* ; Phycoerythrin ; Sciatic Nerve ; Spleen*

This study was purposed to investigate the effect of phycocyanin at different concentration on proliferation of K562 cells, to detect the changes of integrin beta1 expression and intracellular focal adhesion kinase (FAK) gene expression on the surface K562 cells treated with phycocyanin, and to explore the possible mechanism of integrin beta1 effect on phycocyanin inhibiting proliferation of K562 cells. The expression level of integrin beta1 on the surface of K562 cells was evaluated by flow cytometry (FCM); the growth of K562 cells treated with phycocyanin was measured by MTT assay; the expression level of FAK mRNA was analyzed by relatively quantitative RT-PCR after four-day culture of K562 cells with phycocyanin of 40 microg/ml, 80 microg/ml and 160 microg/ml, respectively. The results showed that integrin beta1 expression on the surface of K562 cells was significantly higher than that in bone marrow mononuclear cells (BMMNC) from normal subjects. Phycocyanin could not change the level of integrin beta1 expression. Phycocyanin could increase the expression of FAK gene on K562 cells and inhibit the proliferation of K562 cells. It is concluded that phycocyanin can inhibit the proliferation of K562 cells through enhancing the conjunction of cell stroma with integrin beta1 on K562 cell surface, up-regulating the expression level of FAK gene in K562 cells, restoring the signaling pathway of proliferation inhibition mediated by integrin beta1. The possible mechanism of phycocyanin in the proliferation inhibition of K562 cells is to increase the expression of FAK gene. The phycocyanin may be considered as a potential agent for inhibition of cancer cell proliferation.
Antineoplastic Agents, Phytogenic ; pharmacology ; Cell Proliferation ; drug effects ; Focal Adhesion Kinase 1 ; biosynthesis ; genetics ; Humans ; Integrin beta1 ; biosynthesis ; genetics ; K562 Cells ; Phycocyanin ; pharmacology ; RNA, Messenger ; biosynthesis ; genetics