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Bioactive-Chylomicrons for Oral Lymphatic Targeting of Berberine Chloride: Novel Flow-Blockage Assay in Tissue-Based and Caco-2 Cell Line Models

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ABSTRACT

Purpose

To develop novel bioactive-chylomicrons to solve oral delivery obstacles of Berberine chloride and target the lymphatic system.

Methods

Berberine-loaded bioactive-chylomicrons were prepared and underwent full in vitro characterization. Intestinal permeability was appraised via both non-everted gut sac model and Caco-2 cell model. Furthermore, Bioactive-chylomicrons’ cellular uptake and distribution were examined by laser scanning confocal microscopy. Finally, a novel chylomicron flow-blockage assay on tissue and cellular levels were elaborated to assess the lymphatic targeting ability.

Results

Berberine-loaded chylomicrons showed spherical vesicles of size (175.6 nm), PDI (0.229), zeta potential (−16 .6 mV) and entrapment efficiency (95.5%). Ex-vivo intestinal permeability studies demonstrated 10.5 fold enhancement in permeability of Berberine-loaded chylomicrons over free Berberine. Moreover, Caco-2 studies revealed significant improvement in chylomicrons’ permeability and cellular uptake. Furthermore, confocal microscopy analyses revealed 2 fold increase in berberine-loaded chylomicrons’ intracellular fluorescence. Lymphatic targeting models were successfully elaborated using cycloheximide protein synthesis inhibitor. Such models demonstrated 47 and 27.5% reduction in ex-vivo and Caco-2 permeability respectively. Finally, a good rank order correlation was established between different permeability assessment techniques.

Conclusion

The findings shed the light on the underlying mechanisms of Berberine bioavailability improvement. Consequently, berberine-loaded chylomicrons could be considered as promising bioactive-nanocarriers for Berberine lymphatic targeting and bioavailability improvement.

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Abbreviations

BER:

Berberine chloride

BSA:

Bovine serum albumin

CM:

Chylomicron

CHX:

Cycloheximide

LSCM:

Laser scanning confocal microscopy

P-gp:

p-glycoprotein

PDI:

Polydispersity index

ROI:

Region of interest

CMo :

Unmodified chylomicron (surfactant free)

REFERENCES

  1. Nguyen TX, Huang L, Liu L, Elamin Abdalla AM, Gauthier M, Yang G. Chitosan-coated nano-liposomes for the oral delivery of berberine hydrochloride. J Mater Chem B. 2014;2:7149–59.

    Article  CAS  Google Scholar 

  2. Xue M, Yang MX, Zhang W, Li XM, Gao DH, Ou ZM, et al. Characterization, pharmacokinetics, and hypoglycemic effect of berberine loaded solid lipid nanoparticles. Int J Nanomedicine. 2013;8:4677–87.

    Article  PubMed  PubMed Central  Google Scholar 

  3. Ortiz LMG, Lombardi P, Tillhon M, Scovassi AI. Berberine, an epiphany against cancer. Molecules. 2014;19:12349–67.

    Article  PubMed  Google Scholar 

  4. Goto H, Kariya R, Shimamoto M, Kudo E, Taura M, Katano H, et al. Antitumor effect of berberine against primary effusion lymphoma via inhibition of NF-B pathway. Cancer Sci. 2012;103:775–81.

    Article  CAS  PubMed  Google Scholar 

  5. Zhang Z, Chen Y, Deng J, Jia X, Zhou J, Lv H. Solid dispersion of berberine–phospholipid complex/TPGS 1000/SiO2: preparation, characterization and in vivo studies. Int J Pharm. 2014;465:1–11.

    Article  Google Scholar 

  6. Sailor G, Seth AK, Parmar G, Chauhan S, Javia A. Formulation and in vitro evaluation of berberine containing liposome optimized by 32 full factorial designs. J Appl Pharm Sci. 2015;5:23–8.

    Article  Google Scholar 

  7. Liu Y-T, Hao H-P, Xie H-G, Lai L, Wang Q, Liu C-X, et al. Extensive intestinal first-pass elimination and predominant hepatic distribution of berberine explain its low plasma levels in rats. Drug Metab Dispos. 2010;38:1779–84.

    Article  CAS  PubMed  Google Scholar 

  8. Chen W, Miao Y-Q, Fan D-J, Yang S-S, Lin X, Meng L-K, et al. Bioavailability study of berberine and the enhancing effects of TPGS on intestinal absorption in rats. AAPS PharmSciTech. 2011;12:705–11.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  9. Souza CR, Oliveira HR, Pinheiro WM, Biswaro LS, Azevedo RB, Gomes AJ, et al. Gold nanoparticle and berberine entrapped into hydrogel matrix as drug delivery system. J Biomater Nanobiotechnol. 2015;6:53–63.

    Article  CAS  Google Scholar 

  10. Interaction L. New oral delivery system to improve absorption of berberine: likely interaction of cationized chitosan with PG-P pump. Int J Drug Deliv Technol. 2014;5:33–42.

    Google Scholar 

  11. Fan Z, Wu J, Fang X, Sha X. A new function of vitamin E-TPGS in the intestinal lymphatic transport of lipophilic drugs: enhancing the secretion of chylomicrons. Int J Pharm. 2013;445:141–7.

    Article  CAS  PubMed  Google Scholar 

  12. Ghosh S, Roy T. Nanoparticulate drug-delivery systems: lymphatic uptake and its gastrointestinal applications. J Appl Pharm Sci. 2014;4:130–23.

    Google Scholar 

  13. Van Greevenbroek MMJ, De Bruin TWA. Chylomicron synthesis by intestinal cells in vitro and in vivo. Atherosclerosis. 1998;141:9–16.

    Article  Google Scholar 

  14. Dahan A, Hoffman A. Evaluation of a chylomicron flow blocking approach to investigate the intestinal lymphatic transport of lipophilic drugs. Eur J Pharm Sci. 2005;24:381–8.

    Article  CAS  PubMed  Google Scholar 

  15. Sek L, Porter CJH, Charman WN. Characterisation and quantification of medium chain and long chain triglycerides and their in vitro digestion products, by HPTLC coupled with in situ densitometric analysis. J Pharm Biomed Anal. 2001;25(3–4):651–61.

    Article  CAS  PubMed  Google Scholar 

  16. Gershkovich P, Hoffman A. Uptake of lipophilic drugs by plasma derived isolated chylomicrons: linear correlation with intestinal lymphatic bioavailability. Eur J Pharm Sci. 2005;26:394–404.

    Article  CAS  PubMed  Google Scholar 

  17. Dierling AM, Cui Z. Targeting primaquine into liver using chylomicron emulsions for potential vivax malaria therapy. Int J Pharm. 2005;303:143–52.

    Article  CAS  PubMed  Google Scholar 

  18. Jain V, Nath B, Gupta GK, Shah PP, Siddiqui MA, Pant AB, et al. Galactose-grafted chylomicron-mimicking emulsion: evaluation of specificity against HepG-2 and MCF-7 cell lines. J Pharm Pharmacol. 2009;61:303–10.

    Article  CAS  PubMed  Google Scholar 

  19. Freag MS, Elnaggar YSR, Abdallah OY. Lyophilized phytosomal nanocarriers as platforms for enhanced diosmin delivery: optimization and ex-vivo permeation. Int J Nanomedicine. 2013;8:2385–97.

    PubMed  PubMed Central  Google Scholar 

  20. Bothiraja C, Pawar A, Deshpande G. Ex-vivo absorption study of a nanoparticle based novel drug delivery system of vitamin D3 (Arachitol NanoTM) using everted intestinal sac technique. J Pharm Investig. 2016;46:425–32.

    Article  CAS  Google Scholar 

  21. Lin YC, Kuo JY, Hsu CC, Tsai WC, Li WC, Yu MC, et al. Optimizing manufacture of liposomal berberine with evaluation of its antihepatoma effects in a murine xenograft model. Int J Pharm. 2013;441:381–8.

    Article  CAS  PubMed  Google Scholar 

  22. Hanchinalmath JV, Londonkar R. Cytotoxic and apoptosis-inducing effect of luteolin isolated from feronia limonia on hepg2 cells. Biolife. 2012;2:1287–92.

    Google Scholar 

  23. Tormalehto S, Monkkonen H, Asunmaa K, Niemi R, Auriola S, Vepsalainen J, et al. C ellular uptake and metabolism of clodronate and its derivatives in Caco-2 cells: a possible correlation with bisphosphonate-induced gastrointestinal. Cell Prolif. 2003;19:23–9.

    Google Scholar 

  24. Tang C, Wu XD, Yu YM, Duan H, Zhou J, Xu L. Cell extraction combined with off-line HPLC for screening active compounds from Coptis chinensis. Biomed Chromatogr. 2016;30:658–62.

    Article  CAS  PubMed  Google Scholar 

  25. Sera TL, Paulo W, Vilma JO, Perkins E, Parke D, Holy J. Different concentrations of berberine result in distinct cellular localization patterns and cell cycle effects in a melanoma cell line. Cancer Chemother Pharmacol. 2008;61:1007–18.

    Article  Google Scholar 

  26. Ismail M. The use of Caco-2 cells as an in vitro method to study bioavailability of iron. Mal J Nutr. 1999;5:31–45.

    Google Scholar 

  27. Martel F, Monteiro R, Lemos C. Uptake of serotonin at the apical and basolateral membranes of human intestinal epithelial (Caco-2) cells occurs through the neuronal serotonin transporter (SERT). J Pharmacol Exp Ther. 2003;306:355–62.

    Article  CAS  PubMed  Google Scholar 

  28. Zidan AS, Spinks CB, Habib MJ, Khan MA. Formulation and transport properties of tenofovir loaded liposomes through Caco-2 cell model. J Liposome Res. 2013;2104:1–9.

    Google Scholar 

  29. Field EJ, Albright E, Mathur SN. Regulation of trigiyceride-rich Lipoproteinsecretion by fatty acids in Caco-2 cells. J Lipid Res. 1988;29:1427–37.

    CAS  PubMed  Google Scholar 

  30. Paliwal R, Paliwal SR, Mishra N, Mehta A, Vyas SP. Engineered chylomicron mimicking carrier emulsome for lymph targeted oral delivery of methotrexate. Int J Pharm. 2009;380:181–8.

    Article  CAS  PubMed  Google Scholar 

  31. Reix N, Parat A, Seyfritz E, Van Der Werf R, Epure V, Ebel N, et al. In vitro uptake evaluation in Caco-2 cells and in vivo results in diabetic rats of insulin-loaded PLGA nanoparticles. Int J Pharm. 2012;437:213–20.

    Article  CAS  PubMed  Google Scholar 

  32. Bradford MM. A rapid and sensitive method for the quantitation of microgram quantities of protein utilizing the principle of protein-dye binding. Anal Biochem. 1976;72:248–54.

    Article  CAS  PubMed  Google Scholar 

  33. Pan GY, Wang GJ, Liu XD, Fawcett JP, Xie YY. The involvement of P-glycoprotein in berberine absorption. Pharmacol Toxicol. 2002;91:193–7.

    Article  CAS  PubMed  Google Scholar 

  34. Rege BD, Kao JPY, Polli JE. Effects of nonionic surfactants on membrane transporters in Caco-2 cell monolayers. Eur J Pharm Sci. 2002;16:237–46.

    Article  CAS  PubMed  Google Scholar 

  35. Seeballuck F, Lawless E, Ashford MB, O’Driscoll CM. Stimulation of triglyceride-rich lipoprotein secretion by polysorbate 80: in vitro and in vivo correlation using caco-2 cells and a cannulated rat intestinal lymphatic model. Pharm Res. 2004;21:2320–6.

    Article  CAS  PubMed  Google Scholar 

  36. Artursson P, Palm K, Luthman K. Caco-2 monolayers in experimental and theoretical predictions of drug transport. Adv Drug Deliv Rev. 2012;64:280–9.

    Article  Google Scholar 

  37. Elsheikh MA, Elnaggar YSR, Gohar EY, Abdallah OY. Nanoemulsion liquid preconcentrates for raloxifene hydrochloride: optimization and in vivo appraisal. Int J Nanomedicine. 2012;7:3787–802.

    CAS  PubMed  PubMed Central  Google Scholar 

  38. Yang H-T, Wang G-J. Transport and uptake characteristics of a new derivative of berberine (CPU-86017) by human intestinal epithelial cell line: caco-2. Acta Pharmacol Sin. 2003;24:1185–91.

    CAS  PubMed  Google Scholar 

  39. Trevaskis NL, Kaminskas LM, Porter CJH. From sewer to saviour — targeting the lymphatic system to promote drug exposure and activity. Nat Rev Drug Discov. 2015;14:781–803.

    Article  CAS  PubMed  Google Scholar 

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Acknowledgements

The authors are deeply grateful to Centre of Excellence for Research in Regenerative Medicine and its Applications (CERRMA) at Faculty of Medicine, Alexandria University, Egypt where Cell culture experiments were conducted. The authors also acknowledge the Medical Research Centre I at the Faculty of Medicine, Alexandria University. Egypt where the HPLC analyses were performed.

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Authors and Affiliations

  1. Department of Pharmaceutics, Faculty of Pharmacy, Alexandria University, 1-Azarita Square, Alexandria, Egypt

    Manal A. Elsheikh, Yosra S. R. Elnaggar & Ossama Y. Abdallah

  2. Department of Pharmaceutics, Faculty of Pharmacy, Damanhour University, Damanhour, Egypt

    Manal A. Elsheikh

  3. Department of Pharmaceutics, Faculty of Pharmacy and Drug Manufacturing, Pharos University in Alexandria, Alexandria, Egypt

    Yosra S. R. Elnaggar

  4. Center of Excellent for Research in Regenerative Medicine and its Application (CERRMA), Faculty of Medicine, Alexandria University, Alexandria, Egypt

    Dina Y. Otify

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  1. Manal A. Elsheikh
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  2. Yosra S. R. Elnaggar
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  3. Dina Y. Otify
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  4. Ossama Y. Abdallah
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Correspondence to Yosra S. R. Elnaggar.

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Elsheikh, M.A., Elnaggar, Y.S.R., Otify, D.Y. et al. Bioactive-Chylomicrons for Oral Lymphatic Targeting of Berberine Chloride: Novel Flow-Blockage Assay in Tissue-Based and Caco-2 Cell Line Models. Pharm Res 35, 18 (2018). https://doi.org/10.1007/s11095-017-2307-z

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  • DOI: https://doi.org/10.1007/s11095-017-2307-z

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