Synthesis, characterizations, and biocompatibility evaluation of polycaprolactone–MXene electrospun fibers

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Synthesis, characterizations, and biocompatibility evaluation of polycaprolactone–MXene electrospun fibers Ganesh Prasad Awasthia, Bikendra Maharjana, Sita Shresthaa, Deval Prasad Bhattaraia,b, Deockhee Yoona, Chan Hee Parka,c,*, Cheol Sang Kima,c,* a

Department of Bionanosystem Engineering, Graduate School, Jeonbuk National University, Jeonju, 561-756, Republic of Korea Department of Chemistry, Amrit Campus, Tribhuvan University, Kathmandu, Nepal c Division of Mechanical Design Engineering, Jeonbuk National University, Jeonju, 561-756, Republic of Korea b

G R A P H I C A L A B S T R A C T

A R T I C LE I N FO

A B S T R A C T

Keywords: Polycaprolactone MXene Electrospinning Fibroblasts Pre-osteoblasts Biocompatibility

The two-dimensional (2D) MXene has attracted great interest in the field of biomedical applications. Here, we synthesized for the first time, polycaprolactone-MXene (PCL-MXene) composite electrospun fibers and evaluated its possible applications in biomedical areas. The composite fibers were characterized by field emission scanning electron microscopy, transmission electron microscopy, X-ray diffraction, Fourier transform infrared spectroscopy, X-ray photoelectron spectroscopy, thermogravimetric analysis/differential scanning calorimetry, mechanical strength, and contact angle. Additionally, biocompatibility evaluation of the as-synthesized composite fibers was carried out on fibroblasts (NIH-3T3) and preosteoblasts (MC3T3-E1) cell lines, as a model. Besides these, possible biomineralization activity of the composites fibers was also determined for the harden tissues formation. The results showed that PCL-MXene composite electrospun fibers were cell friendly for both cell lines. However, pre-osteoblast cells exhibited higher cell viability compared to fibroblasts. Considering auspicious results, this work is expected to open the possible pathway for the expanded applications of MXene based composites in biomedical applications.

1. Introduction Over the last few years, inorganic 2D material; MXene has been

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increasing attention for researchers due to its hexagonal structure as similar to graphene [1,2]. The name MXene reflects the removal of the metal “A” from the MAX phase or Mn+1AXn obtained the separate

Corresponding authors at: Department of Bionanosystem Engineering, Graduate School, Jeonbuk National University, Jeonju, 561-756, Republic of Korea. E-mail addresses: [email protected] (C.H. Park), [email protected] (C.S. Kim).

https://doi.org/10.1016/j.colsurfa.2019.124282 Received 22 July 2019; Received in revised form 21 November 2019; Accepted 25 November 2019 Available online 26 November 2019 0927-7757/ © 2019 Published by Elsevier B.V.

Please cite this article as: Ganesh Prasad Awasthi, et al., Colloids and Surfaces A, https://doi.org/10.1016/j.colsurfa.2019.124282

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expanded layers. Where, “M” stands for an early transitional metal (Ti, V, Nb, and Mo), “A” stands for the metal of group 13 and 14 of the periodic table, “X” for carbon or nitrogen, and n = 1, 2 or 3 [3]. The outstanding properties of MXene (i.e. conductivity, hydrophilicity, chemical, and mechanical stability), it has been widely used for various research areas such as energy storage, photocatalysis, wireless communication, shielding, as well as biomedical applications [4–9]. Regarding the biomedical applications, MXene has been widely used for photothermal therapy, biosensor, bioimaging and also poised to improve wearable artificial kidney options [10–14]. Thus, biocompatibility assessment of the MXene based biomaterials is essential for overall biomedical applications. Recently, various MXene incorporated biomaterials have been developed to obtain the desirable physicochemical properties trigger to the practical applications. Zhuang; et al. [15] fabricated the iron oxide/tantalum carbide (Ta4C3) MXene composite via in-situ growth for efficient breast cancer theranostics. The authors found that as-synthesized composite has high photothermal conversion efficiency to complete tumor eradication without reoccurrence. Recently polyethylene glycol(PEG) coated titanium carbide (Ti2C) MXene were reported by Szuplewska et al. [10] for photothermal therapy as a notable NIR induced ability to cancerous cells’ ablation. Moreover, Chen et al. [11] prepared TiO2/Ti3C2 quantum dots composite film through the self-assembly method for photoelectrochemical biosensing, which showed the high stability, sensitivity, and selectivity for Glutathione existing in the biological systems. However, these synthesis methods have some challenges such as deficiency of controllable preparation strategies, morphology, particle size, and cost-effectiveness [16,17]. Electrospinning is the versatile technique to produce the polymeric fibrous scaffolds with a diameter range from a micrometer to nanometer. These fibrous scaffolds can mimic a native extracellular matrix hence they are utilized in various biomedical applications such as; tissue engineering, drug delivery, polymer coating stents, etc. For the fiber production, the natural and synthetic polymers: polycaprolactone (PCL), poly (lactic acid) (PLA), poly (glycolic acid) (PGA), cellulose, chitosan, gelatin, etc. are being widely used [18]. Among these, PCL, a synthetic polymer has excellent mechanical strength, and it is also a biodegradable and biocompatible polymer. The electrospun PCL fibers fabricated via electrospinning shows large surface area to volume ratio, flexible, and high porosity which are the most desirable characteristics (such as; protein adsorption, cell binding, cell proliferation) for biomedical as well as tissue regeneration applications [19–23]. Moreover, in our previous work, it was already reported that the polycaprolactone-cellulose acetate-dextran electrospun composite nano mats showed good biocompatibility and effective wound healing efficacy [24]. Not only the polymeric fibers, the polymeric nanoparticles, and nano scaffolds were also widely used in biomedical applications due to their controlled size, shape, composition, and surface properties, which can directly influence targeted biological applications [25]. Y. Shi et al. synthesized the polymeric fluorescent organic nanoparticles (FONs) through self polymerization of dopamine and polyethyleneimine with desirable biocompatibility, high water dispersibility and strong fluorescence for biological imaging [26]. Similarly, Poly (vinyl alcohol) coated polypyrrole nanoparticles were synthesized and used for effective photothermal therapy by Liu et al. [27]. C. Sharma et al. fabricated chitosan-gelatin-alginate-hydroxyapatite based nano biocomposite scaffold by simple foaming method for bone tissue engineering [28]. Currently, other than single component nanofibers, the composite electrospun nanofibers have been produced by surface modification with the incorporation of filler components to obtain the desirable physicochemical and biological properties such as biocompatibility, hydrophilicity, surface roughness and reactivity of materials by the host and guest materials. The filler components of composite nanofibers were incorporated by in-situ and ex-situ methods. During the in-situ synthesis of composite nanofibers, the precursor of the filler is

introduced with a polymer solution. Whereas, pre-synthesized particles are directly mixed in polymeric solution before electrospinning in the ex-situ method [29]. Various fillers including different types of carbonaceous materials (e.g. Graphene, carbon nanotubes) [30–35], nanoparticles (e.g. polymeric, and metals) [36,37] have been widely used to enhance electrical, chemical, mechanical, and thermal properties by blending with different well-established biopolymers for biomedical applications [38]. Beside them, MXene has been rapidly studied 2D inorganic filler material to modify the surface of electrospun nanofibers for biomedical applications [39]. During the preparation of MXene, the hydrofluoric acid etching process induces the hydrophilic groups (such as OH, O, and F) on their surfaces that ascertain its improved hydrophilic properties which lead towards enhanced biocompatibility of the material. In addition, the formation of MXene from biologically inert metals combined with carbide, nitride, or carbonitrides also supports its cell-friendly behavior during cell adhesion, proliferation and differentiation processes [40,41]. Recently, Mayerberger et al. [39] synthesized electrospun Ti3C2Tz (MXene)/chitosan composite fibers for antibacterial properties towards Gram-negative Escherichia coli (E.coli) and Grampositive Staphylococcus aureus (S. aureus). They concluded that as-synthesized composite fibers are non-toxic and promising material in wound healing applications. Hence, in this study, we report for the first-time fabrication of PCL MXene composite fibers by electrospinning method following the ex-situ filler introducing the process. To realize the potential interest in 2D materials for biomedical fields, as-synthesized composite fibers were evaluated physicochemically and biologically. The enhanced desirable properties (such as; hydrophilicity, protein absorption, cell viability) of composite electrospun fibers could ascertain the potential and promising application towards tissue engineering and regenerative medicine. 2. Materials and methods 2.1. Materials In this study, Polycaprolactone (PCL, Mw = 70,000–90,000) was purchased from Sigma –Aldrich, USA; N, N - Dimethylformamide (DMF) and chloroform (CHCl3) were purchased from Samchun, Korea. Titanium aluminum carbide (MAX-phase) (CARBON, UKRAINE), Hydrofluoric acid (Showa, Japan) were purchased. All materials and solvents were analytical grade and used without any further purification. 2.2. Synthesis of MXene MXene or Ti3C2 was synthesized by the removal of an intermediate element from the MAX phase (Ti3AlC2) by HF etching as reported in the literature [42]. Typically, 2 g of commercial Ti3AlC2 powder was added to 40 mL of 49 % HF or (20 mL/g) in polytetrafluoroethylene container and kept for 24 h at room temperature under constant magnetic stirring. After then, the resulting suspension was initially washed with deionized water by centrifuging (5000 rpm, 5 min.) several times and followed by decantation until the pH of the supernatant approached 6. Finally, the products were washed with ethanol and dried in a vacuum at 80 °C overnight. 2.3. Preparation of electrospinning solutions The different masses of as prepared MXene (0.2, 0.5, 1, and 2 wt.%) were well dispersed into a mixture of DMF and chloroform (1:8 v/v) by a magnetic stirrer for 24 h, separately. 10 wt.% PCL was added into each as prepared MXene dispersion and stirred again until the solution became spinnable. Only the PCL solution was prepared without MXene for comparison. 2

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2.4. Fabrication of PCL-MXene composite electrospun fibers

tetrazolium (MTT, Dojindo Molecular technologies) assay. MTT assay based on the viability of cells, the tetrazolium component of MTT is converted to purple-colored formazan crystals by mitochondrial dehydrogenases. At desired days (1, 3 and 5 days) cell-seeded scaffolds were washed with PBS for twice and 200 μl of 2.5 % of pure phenol-free MTT medium was added and incubated. After 4 h the MTT medium was removed and 200 μl of DMSO was added to dissolve formazan crystals. The optical density of the final solution was measured at a wavelength of 590 nm using an ELISA reader. The resulted were expressed as the percentage cell viability in comparison to the control groups by using the formula; Cell viability = Ai/Ac×100 % (Here; Ai = average absorbance of tested samples and Ac = average absorbance of control samples). All the data are repeated thrice in each experiment. Cell attachment on the fibrous mats was observed after counting for 1, 3 and 5 days by chemical fixation with 3.5 % glutaraldehyde for 1.5 h followed by consecutive increments of the concentration of ethanol series (25, 50, 75, and 95 %) for 20 min. Samples were dried overnight in a laminar flow hood. The cell morphology and attachment mode was determined via SEM.

As prepared electrospinning solution was drawn into the 12 mL plastic syringe connected to metal capillary (diameter =0.51 mm) through a plastic tube. For the electrospinning setup, a digital syringe pump (New Era Pump System, Inc; USA) was used to control the flow rate of 1 mL/h at a 15 kV (applied voltage) and 150 mm tip to collector distance. During electrospinning, fibers were collected to the aluminum sheet attached surface of a ground iron drum by a horizontally moving nozzle at ambient room temperature. 2.5. Physicochemical characterizations Surface morphologies of electrospun fibers were observed by field emission scanning electron microscopy (FESEM, Carl Zeiss, supra 40 V P, Japan) and transmission electron microscopy (TEM; JEM-2010, JEOL, Japan). FESEM samples were sputter-coated with the platinum surface on carbon tape prior to imaging. Fiber diameters were measured using Image J software (NIH, USA). The elemental compositions of the electrospun fibers were confirmed by energy-dispersive X-ray spectroscopy and mapping integrated with FESEM. Fourier transform infrared spectroscopy (FTIR; Perkin Elmer, USA), x-ray diffractometer (XRD; Rigaku, Japan) with Cu Kα (λ =1.540 Å) radiation over Bragg angles ranging from 20 to 80°. Thermogravimetric analysis (TGA) and differential scanning calorimetry (DSC) were analyzed by (TGA, Q50 TA Instruments) under 10−600 °C at 10 °C/min under nitrogen atmosphere. The mechanical properties of the mats were measured via an Instron mechanical tester (LLOYD instruments, LR5K plus, UK) using a dog bone specimen-based from ASTM D882-10. Wettability of electrospun fibers was measured by contact angle analyzer (UNI-CAM.M/get soft, South Korea), the functional states existing on the surface of PCLMXene were detected by X-ray photoelectron spectroscopy (XPS; Kratos, U.K. equipped with monochromatic Al-Kα X-ray excitation source).

2.8. Protein adsorption After five days of cells cultured in 37 °C incubator on different scaffolds, the cells were harvested by trypsin after washing with PBS. The cell samples were centrifuged at 15,000 rpm for 15 min with the addition of protein extraction solution (PRO-PREP™, iNtRON Biotechnology). The supernatant was collected for protein assay. The protein concentration of the solution was determined using a microplate reader at wavelength 620 nm. Bio-Rad Protein Assay (Life science) was used for normalization of protein. 3. Results and discussion 3.1. Physiochemical properties

2.6. Biomineralization test The FESEM of as-synthesized MXene was obtained from the original MAX phase as shown in Fig.1. After removal of the Al layer, 2D sheets of MXene was obtained. The almost removal of the Al layer was confirmed by EDS of MXene. Furthermore, the transitional metal and carbides present in 2D sheets of MXene were confirmed by UV absorption. The UV result showed that the MXene exhibits high absorption in the UV region within the range from 225 to 325 nm which may be due to the bandgap energy of oxidized MXene. A similar result was reported by S. Elumalai too [44]. The surface morphology of PCL and PCL-MXene composite electrospun fibers with different weight percent of MXene is shown in Fig. 2. The composite fibers (Fig. 2b-e) with an increasing amount of MXene showed the different fiber morphology with increasing the fiber diameter compared to PCL fibrous mat (Fig. 2a), which is due to the interconnected MXene flakes along the PCL fiber axis [45]. Beads were observed on electrospun fibers from a solution with an increased MXene concentration above 0.5 wt.%. It might be induced by lower viscoelasticity with an increasing amount of MXene [46]. The histograms of fiber diameter of fibrous mats are shown in Fig. 2(a1-e1). The average fiber diameters were observed gradually increased with the increasing amount of MXene (0.69 μm for PCL fibers, 0.83 μm for 0.2 wt.%, 1.32 μm for 0.5 wt.%, 1.35 μm for 1 wt.% and 1.6 μm for 2 wt.% MXene) in composites. Moreover, the presence of MXene in PCL-MXene composite fibers was confirmed by TEM as shown in Fig. 2(f). The TEM images shows the MXene flakes are clearly observed in PCL-MXene composite fibers. The elemental mapping and EDS result of PCL-MXene composite electrospun fibers was shown in Fig. 3. The spatial distribution of elements with additionally observed Ti confirmed the presence of MXene in the composite fibers.

To observe the role of 2D MXene on the surface of PCL fibers for hard tissue engineering applications, a biomimetic mineralization test was carried out to investigate the calcium and phosphate deposition on the surface of electrospun fibers by SBF incubation. The simulated body fluid (SBF) solution (pH 7.2) was prepared as our previous protocol [43]. Briefly, commercially available Hank’s balanced salt (Aldrich, H2387-1) with magnesium sulfate (0.097 g), anhydrous sodium bicarbonate (0.350 g) and calcium chloride (0.185 g) were dissolved in one liter distilled water. The pristine PCL and PCL-MXene composite electrospun fibrous mats (1.0 cm × 1.0 cm) were incubated into 5 mL SBF solution at 37 °C for two weeks. During the incubation, a fresh SBF solution was changed every 24 h. Two weeks of incubated samples were removed from SBF and then washed with deionized water to remove any loosely attached minerals. The obtained samples were dried at room temperature for FESEM analysis. 2.7. Biocompatibility assay Prior to the experiment, the electrospun fibrous mats were sterilized in UV light for 2 h and transferred to 48-well plates. Sterilized samples were rinsed with phosphate buffer saline (pH = 7.4) and incubated overnight within the humidified atmosphere at 37 °C in relevant culture medium (DMEM for fibroblasts and α-MEM for osteoblasts). 500 μL of cell suspension (1 × 104 cells/well, DMEM/α-MEM supplemented with 10 % FBS and 1 % penicillin-streptomycin) was dispensed onto the surface of membranes and incubated at 37 °C in 5 % CO2 atmosphere for different time intervals. The medium was changed every 2 days. The cytotoxicity of MC3T3-E1 and NIH-3T3 cells on different scaffolds were quantified by using 3-(4, 5 - dimethylthiazole-2- yl)-2, 5-diphenyl 3

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Fig. 1. FESEM image of MX phase (Ti3AlC2) (a), MXene (Ti3C2) (b), EDS and UV absorption of MXene (c, d) respectively.

Fig. 2. FESEM images of pristine PCL electrospun fibers (a), PCL-MXene composite electrospun fibers (0.2, 0.5, 1 and 2 wt.% MXene) (b–e) respectively. The TEM image of composite fiber (f) and fiber diameter distributions corresponding to (a–e) are (a1 - e1) respectively. 4

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Fig. 3. Elemental mapping and EDS (bottom) of PCL-MXene (2 wt.%) composite electrospun fibers.

Fig. 4. XRD of PCL, PCL-MXene composite, and MXene powder (A), FTIR of PCL (a), PCL-MXene composite (b) and MXene powder (c) in (B).

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Fig. 5. TGA, DSC, and stress / Strain curves of PCL and PCL-MXene composite electrospun fibers (A, B, C) respectively.

Fig. 6. Wettability test of PCL and PCL-MXene composite electrospun fibers.

The XRD patterns of PCL, PCL-MXene composite fiber and MXene powder are shown in Fig. 4(A). The semi-crystalline nature of PCL fiber showed three significant diffraction peaks at the diffraction angles

2θ = 21.3°, 22.0° and 23.7° corresponding to the crystal planes (110), (111) and (200) respectively [47]. After the introduction of MXene, two additional peaks at the diffraction angles 2θ = 28.6° and 36.2° 6

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Fig. 7. FESEM images of electrospun fibers incubated into SBF solution for 2 weeks: pristine PCL (a), and PCL-MXene composites electrospun fibers (b) with EDS (bottom) respectively.

of MXene into PCL. Even though, strain percentage of the composite fiber was increased having 0.5 wt.% MXene. So, decreasing the order of mechanical strength of composite fibers might be due to the brittleness of MXene. The wettability of as-synthesized composite electrospun fibers was measured by water contact angle measurement as shown in Fig. 6. The wettability of composites was increased with decreasing contact angle by increasing wt.% of MXene in PCL. Therefore, increasing wettability could be supportive for cell adhesion and proliferation in PCL-MXene composite electrospun fibers. The enhanced wettability of composite mats with an increasing weight percentage of Mxene could be associated with their hydrophilic nature due to its terminated hydroxyl or oxygen moieties [53].

corresponding to the (008) and (111) planes existed in the PCL-MXene composite fiber [2,48]. The MXene peaks existed in the composite were shifted from the angle 26.8° and 35.7° indicating that the well physical interaction between PCL and MXene. The FTIR spectra of PCL, PCL-MXene composite electrospun fiber and MXene powder are shown in Fig. 4(B) (a, b, c) respectively. The characteristic absorption bands at 2947 cm−1 and 2862 cm-1 of pristine PCL mat were assigned the stretching bands of CH2 groups, the absorption band at 1721 cm−1 was due to the stretching of the ester group (C]O), The adsorptions at 1476 cm−1 and 1370 cm−1 belonged to asymmetric deformation and symmetric wagging of CH3 respectively. In addition, the peak existed at 1294 cm−1 and 1165 cm-1 were asymmetric and symmetric stretching of CeOeC. The peak at 1242 cm−1 denoted the CH3 vibrations and 1042 cm−1 belongs to CeO stretching and CH bending. The clearly observed broad adsorption band at 469 cm−1 depicted to Ti-C vibration in the PCL-MXene composite electrospun fiber assured the presence of MXene [49,50]. Furthermore, except Ti-C vibration band, two broad absorption bands at 3400 cm−1 and 1515 cm−1 in pristine MXene revealed the presence of strong external water absorbed on its surface [51]. The thermal behavior of PCL and PCL-MXene composites was evaluated by thermogravimetric analysis (TGA) and differential scanning calorimetry (DSC) as shown in Fig. 5(A, B). The characteristic melting endothermic peak of PCL appeared at 65 ºC and the decomposition of about 364 °C [52]. The melting and decomposition temperatures of the composite fibers were decreased with increasing wt.% of MXene in PCL fibers whereas yield wt.% and thermal stabilities were increased between 450–600 °C. Furthermore, DSC analysis also revealed a similar result about the melting point and decomposition temperature of PCL-MXene composite electrospun fibers. Fig. 5(C) shows the stress/strain curves of PCL and PCL-MXene composites electrospun fibers. The mechanical properties of the PCL-MXene composites electrospun fibers could not increase with an increasing amount

3.2. Biomineralization study The in vitro biomineralization test was used to assess the hydroxyapatite nucleation performance of PCL-MXene composite fibers. PCL fibers are well-known substrate used in a tissue engineering which favors the mineralization. However, the mineralization study of the composite containing MXene has not been assessed. Therefore, PCLMXene composites were studied in order to evaluate the nucleation of hydroxyapatite in the SBF solution. Fig.7. shows the FESEM image of PCL and PCL-MXene composite electrospun fibers after immersing in the SBF solution for 14 days. As expected, the PCL-MXene composite fibers have a successful deposition of calcium phosphate minerals qualitatively and quantitatively. The deposition of calcium ions and phosphate ions could be due to the wettability of MXene [54]. The presence of calcium and phosphorus was confirmed by EDS of SBF treated fibrous scaffolds. The proper molar ratio (∼1.6) of calcium and phosphorus deposition to the surface of PCL–MXene electrospun fibers was found to be in good agreement to the standard value in hydroxyapatite crystals [55]. The result confirmed that the sufficient in-vitro 7

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Fig. 8. Biocompatibility test. (a1 and b1) MTT assay, (a2 and b2) SEM images of NIH-3T3 and MC3T3-E1 cell attachment and proliferation on pristine PCL and PCLMXene fibrous scaffolds. (c) Protein adsorption test result. (d1 and d2) XPS spectra of PCL-MXene composite fibers before and after cell culture for 5 days respectively.

the NIH-3T3 and MC3T3-E1 cell adhesion on these electrospun fibers for 3 and 5 days. SEM images of NIH-3T3 and MC3T1-E1 also revealed that the lower number of cell adhesion into the surface of PCL-MXene composite compared to pristine PCL fibers. The protein adsorption test results are given in Fig. 8c. As in the figure, satisfactory interaction between composite electrospun fibers and cell culture protein can be seen. However, we noticed lower protein adsorption in PCL-MXene composite as compared to pristine PCL fibers which might be due to the thick fiber morphology of the composite fiber [56]. In addition, to confirm the presence of MXene in the composite fibers after certain days of cell culture, the XPS (Fig. 8d1 and 8d2) of the PCL-MXene composite scaffold was recorded before and after cell culture. Before cell culture (Fig. 8d1), the survey spectrum showed the coexistence of titanium (Ti), carbon (C), and oxygen (O) on the surface of PCL-MXene composite electrospun fibers corresponding to the binding energy 460.9, 285.1 and 532.4 eV, respectively. After 5 days of cell culture (Fig. 8d2), some additional peaks of Na, Cl and N were noticed, which were corresponding to the binding energy 1069.2, 196.9 and 399.02 eV along with the peak sift of Ti from 460.9–467 eV. The peak sifts of Ti due to the oxidation of Ti at the ambient condition of cell culture media [57].

biomineralization and calcium/phosphorus deposition on the scaffolds are bioactive enough for osteointegration. 3.3. Biocompatibility and cell adhesion study In order to evaluate the cellular response of PCL-MXene composite electrospun fibers, the fibroblast (NIH-3T3) and pre-osteoblast (MC3T3E1) cell were seeded for 1, 3, and 5 days. The biocompatibility and cell adhesion on fibrous scaffolds were determined by using the MTT assay and SEM imaging. The cells seeded Tissue Culture Plate (TCP-48) without electrospun fibers was set as a control group. Fig. 8a1 shows the cell viability on different electrospun fibers after 1, 3, and 5 days cell culture of fibroblasts (NIH-3T3). The NIH-3T3 viability on PCL and PCL-MXene composite fibers was lower than that of control for 1 day. However, cell viability was decreased with an increasing amount of MXene in PCL corresponding to the increasing days of cell culture. In the 5th day’s cell culture of NIH-3T3, cell viability was about ∼70 % compared to the control sample Fig. 8a1. Similarly, the cell viability of preosteoblasts (MC3T3-E1) cells on the electrospun fibers in the same days of cell culture is shown in Fig. 8b1. In the case of MC3T3-E1, cell viability on PCL-MXene composite fibers was also decreased compared to control and PCL sample. However, pre-osteoblast cell viability on the PCL-MXene composite (0.2 and 0.5 w%) showed more (∼72 %) than fibroblast cells. Fig. 8a2 and 8b2) shows the respective SEM images of 8

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4. Conclusions

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