Prodigiosin isolated from Serratia marcescens in the Periplaneta americana gut and its apoptosis‑inducing activity in HeLa cells

Lin,Pei‑Bin; Shen,Juan; Ou,Pei‑Yu; Liu,Ling‑Yan; Chen,Zhi‑Yu; Chu,Fu‑Jiang; Wang,Jie; Jin,Xiao‑Bao

doi:10.3892/or.2019.7089

Prodigiosin isolated from Serratia marcescens in the Periplaneta americana gut and its apoptosis‑inducing activity in HeLa cells

Authors:
- Pei‑Bin Lin
- Juan Shen
- Pei‑Yu Ou
- Ling‑Yan Liu
- Zhi‑Yu Chen
- Fu‑Jiang Chu
- Jie Wang
- Xiao‑Bao Jin
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Affiliations: School of Basic Courses, Guangdong Pharmaceutical University, Guangzhou Higher Education Mega Center, Guangzhou, Guangdong 510006, P.R. China, Guangdong Key Laboratory of Pharmaceutical Bioactive Substances, Guangdong Pharmaceutical University, Guangzhou, Guangdong 510006, P.R. China
Published online on: April 2, 2019 https://doi.org/10.3892/or.2019.7089
Pages: 3377-3385

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Abstract

Serratia marcescens are considered to be abundant and optimal resources for obtaining prodigiosin, which can be isolated from soil, water, plants and air but rarely from insects. In the present study, a strain of Serratia marcescens named WA12‑1‑18 was isolated from the gut of Periplaneta americana, which was capable of producing high levels of pigment reaching 2.77 g/l via solid fermentation and was identified as prodigiosin by ultraviolet, high performance liquid chromatography (LC), Fourier‑transform infrared spectroscopy, LC‑mass spectroscopy and nuclear magnetic resonance. The apoptotic tumor cells treated with prodigiosin were examined by 4',6‑diamidino‑2‑phenylindole (DAPI) staining assays and transmission electron microscopy. Flow cytometry (FCM) was utilized to measure the apoptotic rate with Annexin V staining and the expression levels of proteins involved in apoptosis, including B‑cell lymphoma 2 (Bcl‑2), Bcl‑2‑associated X (Bax) and caspase‑3 were determined by western blot analysis and reverse transcription‑quantitative polymerase chain reaction (RT‑qPCR). The experimental results revealed that prodigiosin could inhibit the proliferation of HeLa cells and the half‑maximal inhibitory concentration values of prodigiosin in HeLa were 2.1, 1.2 and 0.5 µg/ml over 24, 48 and 72 h, respectively. Furthermore, DAPI staining assays and transmission electron microscopy clearly demonstrated that prodigiosin could induce HeLa cell apoptosis. FCM results revealed that the cell apoptotic rates were 19.7±1.4, 23.7±2.4 and 26.2±2.3% following the treatment with 0.5, 1.0 and 2.0 µg/ml prodigiosin for 48 h, respectively. Western blot analysis and RT‑qPCR revealed that prodigiosin could activate apoptosis‑associated molecules including Bcl‑2, Bax and caspase‑3. Therefore, the results of the present study demonstrated that the prodigiosin could induce apoptosis in HeLa cells, which may be associated with the upregulation of Bax and caspase‑3, the concomitant downregulation of Bcl‑2 levels and also triggering the extrinsic apoptotic signaling pathway.

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1	Elkenawy NM, Yassin AS, Elhifnawy HN and Amin MA: Optimization of prodigiosin production by Serratia marcescens using crude glycerol and enhancing production using gamma radiation. Biotechnol Rep. 14:47–53. 2017. View Article : Google Scholar
2	Zhang HC, Geng FN and Shen YM: The insect gut microbiota plays crucial roles in modulating the interactions between the host and intestinal pathogens. Chin J Ethnomed Ethnopharmacy. 26:57–60. 2016.
3	Wang J: Effect of Periplaneta americana extract on H125 lung cancer cells. Chin J Public Health. 30:1400–1402. 2014.
4	Wang J and Xin L: Effects of Periplaneta americana L. Extract on human hepatoma cells Bel-7402. Chin J Modern Appl Pharm. 29:876–880. 2012.
5	Oh DC, Kauffman CA, Jensen PR and Fenical W: Induced production of emericellamides A and B from the marine-derived fungus emericella sp. in competing co-culture. J Nat Prod. 70:515–520. 2007. View Article : Google Scholar : PubMed/NCBI
6	Rastogi S, Zhang D and Davis JT: The natural product prodigiosin binds G-quadruplex DNA. Supramolecular Chem. 28:1–12. 2016. View Article : Google Scholar
7	Xia Y, Wang G, Lin X, Song X and Ai L: Solid-state fermentation with Serratia marcescens, Xd-1 enhanced production of prodigiosin by using bagasse as an inertia matrix. Annal Microbiol. 66:1239–1247. 2016. View Article : Google Scholar
8	Nakashima T, Tamura T, Kurachi M, Yamaguchi K and Oda T: Apoptosis mediated cytotoxicity of prodigiosin-like red pigment produced by gamma proteobacterium and its multiple bioactivities. Biol Pharm Bull. 28:2289–2295. 2005. View Article : Google Scholar : PubMed/NCBI
9	Sumathi C, MohanaPriya D, Swarnalatha S, Dinesh MG and Sekaran G: Production of prodigiosin using tannery fleshing and evaluating its pharmacological effects. ScientificWorldJournal. 2014:290–327. 2014. View Article : Google Scholar
10	Osei-Poku J, Mbogo CM, Palmer WJ and Jiggins FM: Deep sequencing reveals extensive variation in the gut microbiota of wild mosquitoes from Kenya. Mol Ecol. 21:5138–5150. 2012. View Article : Google Scholar : PubMed/NCBI
11	Suzuki N, Ohtaguro N, Yoshida Y, Hirai M, Matsuo H, Yamada YL, Imamura N and Tsuchiya T: A compound inhibits biofilm formation of staphylococcus aureus from streptomyces. Biol Pharm Bull. 38:889–892. 2015. View Article : Google Scholar : PubMed/NCBI
12	Douglas AE: Multiorganismal insects: Diversity and function of resident microorganisms. Annu Rev Entomol. 60:17–34. 2015. View Article : Google Scholar : PubMed/NCBI
13	Someya N, Nakajima M, Hamamoto H, Yamaguchi I and Akutsu K: Effects of light conditions on prodigiosin stability in the biocontrol bacterium Serratia marcescens, strain b2. J General Plant Pathol. 70:367–370. 2004. View Article : Google Scholar
14	Hassankhani R, Sam MR, Esmaeilou M and Ahangar P: Prodigiosin isolated from cell wall of Serratia marcescens alters expression of apoptosis-related genes and increases apoptosis in colorectal cancer cells. Med Oncol. 32:3662015. View Article : Google Scholar : PubMed/NCBI
15	Miao L, Wang X, Jiang W, Yang S, Zhou H, Zhai Y, Zhou X and Dong K: Optimization of the culture condition for an antitumor bacterium serratia proteamacula 657 and identification of the active compounds. World J Microbiol Biotechnol. 29:855–863. 2013. View Article : Google Scholar : PubMed/NCBI
16	Patil CD, Patil SV, Salunke BK and Salunkhe RB: Prodigiosin produced by Serratia marcescens NMCC46 as a mosquito larvicidal agent against Aedes aegypti and Anopheles stephensi. Parasitol Res. 109:1179–1187. 2011. View Article : Google Scholar : PubMed/NCBI
17	Marsac P, Pierard N, Porot L, Wim Van den bergh, Grenfell J, Mouillet V, Pouget S, Besamusca J, Farcas F, Gabet T, et al: Potential and limits of FTIR methods for reclaimed asphalt characterization. Materials Structures. 47:1273–1286. 2014. View Article : Google Scholar
18	Pradeep S, Sarath Josh MK, Balachandran S, Sudha Devi R, Sadasivam R, Thirugnanam PE, Doble M, Anderson RC and Benjamin S: Achromobacter denitrificans, SP1 produces pharmaceutically active 25C prodigiosin upon utilizing hazardous di(2-ethylhexyl)phthalate. Bioresour Technol. 171:482–486. 2014. View Article : Google Scholar : PubMed/NCBI
19	Danevčič T, Borić Vezjak M, Tabor M, Zorec M and Stopar D: Prodigiosin induces autolysins in actively grown bacillus subtilis cells. Front Microbiol. 7:272016. View Article : Google Scholar : PubMed/NCBI
20	Kimyon Ö, Das T, Ibugo AI, Kutty SK, Ho KK, Tebben J, Kumar N and Manefield M: Serratia secondary metabolite prodigiosin inhibits Pseudomonas aeruginosa biofilm development by producing reactive oxygen species that damage biological molecules. Front Microbiol. 7:9722016. View Article : Google Scholar : PubMed/NCBI
21	Darshan N and Manonmani HK: Prodigiosin inhibits motility and activates bacterial cell death revealing molecular biomarkers of programmed cell death. AMB Express. 6:502016. View Article : Google Scholar : PubMed/NCBI
22	Lee DJ, Lee JB, Jang HA, Ferrandon D and Lee BL: An antimicrobial protein of the riptortus pedestris, salivary gland was cleaved by a virulence factor of Serratia marcescens. Dev Comp Immunol. 67:427–443. 2017. View Article : Google Scholar : PubMed/NCBI
23	Chen W, Liu Y, Zhang L, Gu X, Liu G, Shahid M, Gao J, Ali T and Han B: Nocardia cyriacigeogica from bovine mastitis induced in vitro apoptosis of bovine mammary epithelial cells via activation of mitochondrial-caspase pathway. Front Cell Infect Microbiol. 7:1942017. View Article : Google Scholar : PubMed/NCBI
24	Zhang J, Liu H, Sun Z, Xie J, Zhong G and Yi X: Azadirachtin induced apoptosis in the prothoracic gland in Bombyx mori and a pronounced Ca²⁺ release effect in Sf9 cells. Int J Biol Sci. 13:1532–1539. 2017. View Article : Google Scholar : PubMed/NCBI
25	Zheng B, Zhou Y, Zhang H, Yang G, Hong Z, Han D, Wang Q, He Z, Liu Y, Wu F, et al: Dl-3-n-butylphthalide prevents the disruption of blood-spinal cord barrier via inhibiting endoplasmic reticulum stress following spinal cord injury. Int J Biol Sci. 13:1520–1532. 2017. View Article : Google Scholar : PubMed/NCBI
26	Livak KJ and Schmittgen TD: Analysis of relative gene expression data using real-time quantitative PCR and the 2^−ΔΔCT method. Methods. 25:402–408. 2001. View Article : Google Scholar : PubMed/NCBI
27	Demain AL and Fang A: The natural functions of secondary metabolites. Adv Biochem Eng Biotechnol. 69:1–39. 2000.PubMed/NCBI
28	Kan SC, Zang CZ, Yeh CW, Chang WF, Lin CC, Hung TH, Shieh CJ and Liu YC: Enhanced bioconversion rate and released substrate inhibition in (R)-phenylephrine whole-cell bioconversion via partial acetone treatment. Enzyme Microb Technol. 86:34–38. 2016. View Article : Google Scholar : PubMed/NCBI
29	Sruthy PB, Anjana JC, Rathinamala J and Jayashree S: The role of red pigment prodigiosin from bacteria of earthworm gut as an anticancer agent. J Microbiol Biotechnol Food Sci. 4:246–251. 2015. View Article : Google Scholar
30	Lapenda JC, Silva PA, Vicalvi MC, Sena KX and Nascimento SC: Antimicrobial activity of prodigiosin isolated from Serratia marcescens UFPEDA 398. World J Microbiol Biotechnol. 31:399–406. 2015. View Article : Google Scholar : PubMed/NCBI
31	Mienda BS, Idi A and Umar A: Microbiological features of solid state fermentation and its applications - An overview. Res Biotechnol. 2:21–26. 2011.
32	Kurbanoglu EB, Ozdal M, Ozdal OG and Algur OF: Enhanced production of prodigiosin by Serratia marcescens mo-1 using ram horn peptone. Braz J Microbial. 46:631–637. 2015. View Article : Google Scholar
33	Wei YH and Chen WC: Enhanced production of prodigiosin-like pigment from Serratia marcescens, SMR by medium improvement and oil-supplementation strategies. J Biosci Bioeng. 99:616–622. 2005. View Article : Google Scholar : PubMed/NCBI
34	Suryavanshi MV, Waghmode SR, Bharti N, Choudhari PB, Hingamire TB and Shouche YS: Isolation and virtual screening of antimicrobial prodigiosin pigment from oxalotrophic Serratia marcescens OX_R strain. J Appl Pharm Sci. 6:52–58. 2016. View Article : Google Scholar
35	Su JC, Chang JH, Huang JW, Chen PP, Chen KF, Tseng PH and Shiau CW: Copper-obatoclax derivative complexes mediate DNA cleavage and exhibit anti-cancer effects in hepatocellular carcinoma. Chem Biol Interact. 228:108–113. 2015. View Article : Google Scholar : PubMed/NCBI
36	Zhou W, Jin ZX and Wan YJ: Apoptosis of human lung adenocarcinoma A549 cells induced by prodigiosin analogue obtained from an entomopathogenic bacterium Serratia marcescens. Appl Microbiol Biotechnol. 88:1269–1275. 2010. View Article : Google Scholar : PubMed/NCBI
37	Pérez-Tomás R and Vinas M: New insights on the antitumoral properties of prodiginines. Curr Med Chem. 17:2222–2231. 2010. View Article : Google Scholar : PubMed/NCBI
38	Montaner B, Navarro S, Piqué M, Vilaseca M, Martinell M, Giralt E, Gil J and Pérez-Tomás R: Prodigiosin from the supernatant of Serratia marcescens induces apoptosis in haematopoietic cancer cell lines. Br J Pharmacol. 131:585–593. 2000. View Article : Google Scholar : PubMed/NCBI
39	Elahian F, Moghimi B, Dinmohammadi F, Ghamghami M, Hamidi M and Mirzaei SA: The anticancer agent prodigiosin is not a multidrug resistance protein substrate. DNA Cell Biol. 32:90–97. 2013. View Article : Google Scholar : PubMed/NCBI