Osteogenesis-promoting Effects of the Electrospun Nanofibers Containing Decellularized Bone Matrix †

doi:10.7503/cjcu20190524

Abstract

Abstract:

Porcine decellularized bone matrix(DBM) was firstly prepared, and then solubilized via pepsin digestion. Solubilized DBM and poly(L-lactic acid)(PLLA) were subsequently processed into PLLA/DBM electrospun nanofibers through eletrospinning technology. The morphology, wettability and mineralization ability of PLLA/DBM electrospun nanofibers were examined, and cytocompatibility and osteogenic differentiation capacity were evaluated by culturing bone marrow mesenchymal cells(BMSCs) on PLLA/DBM electrospun nanofibers. The results showed that cellular components in cancellous bone could be effectively removed after decellularization, as evidenced by the significant decrease in DNA content. The solubilized DBM could be dissolved in 1,1,1,3,3,3-hexafluoro-2-propanol(HFIP) for electrospinning of the PLLA/DBM composite nanofibers formulated at different mass ratios(10∶0, 9∶1, 7∶3 and 5∶5). The obtained PLLA/DBM electrospun nanofibers were demonstrated to be hydrophilic and cytocompatible, and significantly promoted the attachment and osteogenic differentiation of the BMSCs. Furthermore, the presence of DBM facilitated mineral deposition on the nanofiber surface in simulated body fluid(SBF), which suggests its great potential in promoting bonding with the natural bone tissue.

Key words: Extracellular matrix, Decellularized bone matrix, Electrospinning, Bone tissue engineering

CLC Number:

O636

TrendMD:

QIN Chunping, WANG Xianliu, TANG Han, YI Bingcheng, LIU Chang, ZHANG Yanzhong. Osteogenesis-promoting Effects of the Electrospun Nanofibers Containing Decellularized Bone Matrix ^†[J]. Chem. J. Chinese Universities, 2020, 41(4): 780.

Figures/Tables 9

Fig.1 Optical(A1, A2), HE staining(B1, B2) and Masson trichrome staining(C1, C2) images of demineralized bone matrix(A1—C1) and decellularized bone matrix(A2—C2)

Fig.2 DNA content comparison between decellularized bone matrix and the native bone(**P< 0.01)

Fig.3 SEM images(A—D) and the corresponding diameter distributions(E—H) of PLLA/DBM-10/0(A, E), PLLA/DBM-9/1(B, F), PLLA/DBM-7/3(C, G) and PLLA/DBM-5/5(D, H) electrospun nanofibers

Fig.4 FTIR spectra(A) and water contact angles(B) of PLLA/DBM electrospun nanofibers (A) a. DBM; b. PLLA/DBM-10/0; c. PLLA/DBM-9/1; d. PLLA/DBM-7/3; e. PLLA/DBM-5/5. (B) a. PLLA/DBM-10/0; b. PLLA/DBM-9/1; c. PLLA/DBM-7/3; d. PLLA/DBM-5/5.

Fig.5 Cell proliferation of BMSCs on PLLA/DBM electrospun nanofibersa. PLLA/DBM-10/0; b. PLLA/DBM-9/1; c. PLLA/DBM-7/3; d. PLLA/DBM-5/5; e. TCP. *P< 0.05, **P< 0.01.

Fig.6 Cell morphologies of BMSCs cultured on PLLA/DBM-10/0(A, C) and PLLA/DBM-7/3(B, D) electrospun nanofibers after 2 h(A, B) and 24 h(C, D)

Fig.7 ALP staining(A, B), quantified ALP activity(C, **P<0.01), ARS staining(D, E) and ARS quantification(F, **P<0.01) of BMSCs cultured on PLLA/DBM electrospun nanofibers(A), (D) PLLA/DBM-10/0; (B), (E) PLLA/DBM-7/3.

Fig.8 SEM images(A—D) and XRD patterns(E) of PLLA/DBM electrospun nanofibers after being incubated in SBF(A) PLLA/DBM-10/0, 7 d; (B) PLLA/DBM-7/3, 7 d; (C) PLLA/DBM-10/0, 14 d; (D) PLLA/DBM-7/3, 14 d. (E) a. PLLA/DBM-7/3; b. PLLA/DBM-7/3 after mineralization for 14 d; c. PLLA/DBM-10/0; d. PLLA/DBM-10/0 after mineralization for 14 d.

Sample	Atomic fraction(%)
Sample	C	O	P	Ca	Mg
PLLA/DBM-10/0	67.69	32.27	0.01	0.03	0
PLLA/DBM-7/3	17.66	52.69	10.43	18.60	0.62

References 30

[1]	Tang D., Tare R. S., Yang L. Y., Williams D. F., Ou K. L., Oreffo R. O. C ., Biomaterials, 2016, 83, 363— 382
[2]	Roseti L., Parisi V., Petretta M., Cavallo C., Desando G., Bartolotti I., Grigolo B., Mater. Sci. Eng. C, 2017, 78, 1246— 1262
[3]	Papadimitropoulos A., Scotti C., Bourgine P., Scherberich A., Martin I ., Bone, 2015, 70, 66— 72
[4]	Bose S., Roy M., Bandyopadhyay A., Trends Biotechnol., 2012, 30( 10), 546— 554
[5]	Ghassemi T., Shahroodi A., Ebrahimzadeh M. H., Mousavian A., Movaffagh J., Moradi A., Arch. Bone Jt. Surg., 2018, 6( 2), 90— 99
[6]	Lee D. J., Diachina S., Lee Y. T., Zhao L., Zou R., Tang N., Han H., Chen X., Ko C. C., J. Tissue Eng., 2016, 7, 1— 11
[7]	Gerhardt L. C., Widdows K. L., Erol M. M., Nandakumar A., Roqan I. S., Ansari T., Boccaccini A. R ., J. Biomed. Mater. Res., Part A, 2013, 101( 3), 827— 841
[8]	Badylak S. F ., Biomaterials, 2007, 28( 25), 3587— 3593
[9]	Rothrauff B. B., Yang G., Tuan R. S., Stem Cell Res. Ther., 2017, 8, 133
[10]	Shekaran A., Garcia A. J ., J. Biomed. Mater. Res., Part A, 2011, 96A( 1), 261— 272
[11]	Hashimoto Y., Funamoto S., Kimura T., Nam K., Fujisato T., Kishida A ., Biomaterials, 2011, 32( 29), 7060— 7067
[12]	Chen G., Lv Y., Methods Mol. Biol., 2017, 1577, 239— 254
[13]	Gruskin E., Doll B. A., Futrell F. W., Schmitz J. P., Hollinger J. O., Adv. Drug Delivery Rev., 2012, 64( 12), 1063— 1077
[14]	Sawkins M. J., Bowen W., Dhadda P., Markides H., Sidney L. E., Taylor A. J., Rose F. R. A. J., Badylak S. F., Shakeshehh K. M., White L. J., Acta Biomater., 2013, 9( 8), 7865— 7873
[15]	Gibson M., Beachley V., Coburn J., Bandinelli P A .,Mao H. Q., Elisseeff J., Bio. Med Res. Int., 2014,469120
[16]	Jose M. V., Thomas V., Johnson K. T., Dean D. R., Nyalro E., Acta Biomater., 2009, 5( 1), 305— 315
[17]	Yao Q., Cosme J. G. L., Xu T., Miszuk J. M., Picciani P. H. S., Fong H., Sun H ., Biomaterials, 2017, 115, 115— 127
[18]	Jang J. H., Castano O., Kim H. W., Adv. Drug Delivery Rev., 2009, 61( 12), 1065— 1083
[19]	Kesireddy V., Kasper F. K., J. Mater. Chem. B, 2016, 4( 42), 6773— 6786
[20]	Gao S., Guo W., Chen M., Yuan Z., Wang M., Zhang Y., Liu S., Xi T., Guo Q., J. Mater. Chem. B, 2017, 5( 12), 2273— 2285
[21]	Yi B. C., Zhang H. L., Yu Z. P., Yuan H. H., Wang X. L., Sheng Y. B., Bao J. Y., Lou X. X., Zhang Y. Z ., Chin. J. Tissue Eng. Res., 2016, 20 52), 7788— 7795
	( 易兵成,张会兰,余哲泡,袁卉华,王先流,沈炎冰,包佳煜,娄向新,张彦中.中国组织工程研究, 2016, 20(52), 7788— 7795)
[22]	Guo J. G., Zhao R., Hou G. Y., Wang F. X ., The Journal of Traditional Chinese Orthopedics and Traumatology, 2001, 13 11), 5— 6
	( 郭建刚,赵然,侯桂英,王芳轩.中医正骨, 2001, 13(11), 5— 6)
[23]	Crapo P. M., Gilbert T. W., Badylak S. F ., Biomaterials, 2011, 32( 12), 3233— 3243
[24]	Feng B., Tu H., Yuan H., Peng H., Zhang Y ., Biomacromolecules, 2012, 13( 12), 3917— 3925
[25]	Wang Y. Q., Cai J. Y., Curr. Appl. Phys., 2007, 7( s1), 108— 111
[26]	Wang L., Wang H., Zhang Z. R., Yin L. H., Yu Z. H., New Chemical Materials, 2014, 42(5), 2391—2397, 129
	( 王琳, 王红, 张忠芮, 殷丽华, 余占海 . 化工新型材料, 2014, 42(5), 46— 48, 129)
[27]	Shih Y. R. V., Chen C. N., Tsai S. W., Wang Y. J., Lee O. K., Stem Cells, 2006, 24( 11), 2391— 2397
[28]	Pietrzak W. S., Ali S. N., Chitturi D., Jacob M., Woodell May J. E., Cell Tissue Banking, 2011, 12( 2), 81— 88
[29]	Kokubo T., Takadama H ., Biomaterials, 2006, 27( 15), 2907— 2915
[30]	Scharnweber D., Born R., Flade K., Roessler S., Stoelzel M., Worch H ., Biomaterials, 2004, 25( 12), 2371— 2380