In Vivo Study of Bone Growth Around Additively Manufactured Implants With Ti‐6Al‐4V and Bioactive Glass Powder Composites

ABSTRACT

Effective osseointegration is a fundamental requirement in biomedical implant applications. Additive manufacturing allows precise control over implant geometry and material composition, enhancing implant design flexibility. Bioactive glass (BG) can substantially enhance bone binding and bioactivity; however, limited research has been conducted on its incorporation into additively manufactured implants. The performance of BG varies depending on the incorporation method, and the spatial and temporal evolution of its integration remains unclear. In this study, we synthesized Ti‐6Al‐4V/58S BG composites by using the selective laser melting method and systematically compared the effects of BG coating and doping in additively manufactured implants. In vivo histological results from animal tests were statistically analyzed and discussed in terms of osseointegration over 4‐ and 12‐week periods. Bone‐to‐implant contact (BIC) and bone density (BD) were used as quantitative metrics to evaluate interactions between the implants and surrounding bone. Our findings indicate that both BG‐doped and BG‐coated implants accelerated bone ingrowth during the early stages of healing. BG‐coated implants demonstrated a greater improvement than did pure 3D‐printed Ti‐6Al‐4V implants. However, the effects of BG became nonsignificant during the later healing stage (12 weeks). This study provides a foundation for systematically investigating BG incorporation methods in 3D‐printed biomedical implants and their effect on osseointegration.

1. Introduction

Implant materials play a pivotal role in the success of orthopedic and dental surgeries [1, 2, 3]. Among these materials, titanium alloys, especially Ti‐6Al‐4V, are widely known for their excellent biocompatibility, corrosion resistance, and mechanical strength [4, 5, 6]. However, the biomechanical mismatch between titanium implants and surrounding tissues remains a problem [7, 8]. Several studies have indicated that differences in stiffness and elastic modulus lead to stress‐shielding, which impairs bone healing and remodeling [9, 10]. Therefore, researchers have begun to focus on developing bone‐like materials through structural modifications [11] and compositional adjustments [12] to mitigate stress‐shielding and promote osseointegration.

Bioactive glasses (BGs) and BG composite materials, which were first introduced by Hench et al. [13, 14], have attracted considerable attention in tissue engineering and drug delivery [15, 16, 17, 18] because of their excellent bioactivity, degradability, and low inflammatory response. BGs form hydroxyapatite (HA) layers on implants and release ion dissolution products, establishing a robust interface between hard and soft tissues and promoting bone growth [19, 20, 21]. Therefore, extensive research has been conducted on BG‐coated layers [22, 23, 24, 25], with a focus on enhancing implant osseointegration performance. Incorporating BG composites into pure metallic biomaterials could improve bone binding and bioactivity. However, the granularity of BGs limits their reliability as space‐making devices. In addition, BGs lack osteoinductive properties and cannot induce bone formation at ectopic sites [26].

Among BGs, 58S exhibits superior apatite formation, bone remodeling and osteoblastic differentiation [27], making it ideal for biomedical applications. In particular, sol‐gel‐derived 58S BGs are considered a promising alternative to glass‐melt‐derived 45S5 BGs because of their superior capacity to induce osteoblastic differentiation [28]. Fathi et al. [29, 30] have demonstrated that 58S BG‐coated 316 L stainless steel improved biocompatibility, leading to earlier implant stabilization and decreased healing time. Our previous study [31] indicated that the spray‐drying method for preparing 58S BGs reduced contamination and improve production efficiency compared with the conventional glass‐melting and sol‐gel methods. In addition, 58S BGs exhibit better bioactivity and a faster HA growth rate than BGs with other compositions do [32]. Thus, 58S BGs are particularly suitable for biomedical applications and were selected for use in the current study.

Additive manufacturing (AM) technology has been widely applied to both ceramics and alloys to achieve complex geometries [11, 33, 34, 35] and has been used to produce BG composite materials. Lam et al. [31] performed histological and histomorphometric analyses to evaluate the properties and in vivo performance of titanium (Ti)‐based alloy and BG composite materials fabricated using the selective laser melting (SLM) method. While BG‐coated and BG‐doped implants have been explored, their comparative in vivo performance remains unclear. This study fills this gap by investigating the chemical and physical interactions of BGs and their temporal and spatial evolution in clinical applications. In this study, we investigated the effect of 0.25 wt% 58S BG applied as both a coating and a dopant in Ti‐6Al‐4V implants made by SLM. Animal implantation tests were conducted using rabbits. Histological results were analyzed using one‐way analysis of variance (ANOVA) to evaluate the performance over short and long periods. This study systematically evaluates the osseointegration performance of Ti‐6Al‐4V implants incorporating 58S BG as a coating and as a dopant. By assessing bone‐to‐implant contact (BIC) and bone density (BD) in an in vivo model, we provide quantitative insights into the effects of BG incorporation on early and long‐term bone healing.

2. Materials and Methods

2.1. Preparation of Implants

In this study, three types of implants were evaluated: Implant I (pure Ti‐6Al‐4V), Implant II (BG‐coated Ti‐6Al‐4V), and Implant III (BG‐doped Ti‐6Al‐4V), as shown in Figure 1a–c, respectively. All implants were fabricated using the same AM process performed at the Industrial Technology Research Institute (ITRI) in Taiwan. This process involved the use of an SLM machine (ITRI‐AM100, Tainan City, Taiwan) with a laser power of 170 W and a scan speed of 1250 mm/s. The implants were designed to replicate the geometry of a commercial Ti‐6Al‐4V implant, ITI 033.512S (Straumann, Basel, Switzerland), with a diameter of 3.3 mm and a length of 10 mm. Ti‐6Al‐4V commercial powder (Titanium Ti64ELI, EOS, Germany) with a particle size of 15–45 µm was used as the base material in this study. In addition, 58S BG powder was selected for its excellent bioactivity.

Figure 1.

SEM images of (a) Implants I (pure Ti‐6Al‐4V), (b)) Implant II (BG‐ coated Ti‐6Al‐4V) X (c) Implant III (BG‐doped Ti‐6Al‐4V) and (d) the surface of Implant II and (e) BG‐doped Ti‐6Al‐4V powder used for Implant III. Implant photos are presented as insets in the corresponding images.

Implant I was fabricated and designated as the control group to investigate the effect of the compositional addition of BG. Implant II was prepared by immersing Implant I in a BG suspension for 10 min, followed by drying in an oven at 70°C for 12 h. The BG coating suspension was prepared by adding 2.5 g of 58S BG to 95.5 g of deionized (DI) water along with 2.0 g of 95 wt% Type I collagen binder (Horien, Taichung, Taiwan). The mixture was stirred at room temperature for 4 h. Implant III was manufactured using BG‐doped Ti‐6Al‐4V powder, as shown in Figure 1d. The Ti‐6Al‐4V commercial powder was doped with 0.25 wt% 58S BG powder that was synthesized as described in our previous study [32]. Specifically, 0.25 g of 58S BG powder and 999.75 g of Ti‐6Al‐4V commercial powder were added to 120 mL of DI water and stirred at 100°C for 3 h. The resulting mixture was dried in an oven at 70°C for 24 h. The concentration of 0.25 wt% was selected because it enabled the use of a flow time of less than 40 s, which was deemed suitable for achieving optimal mechanical properties for 3D printing, as determined in our previous study [31]. Although the mechanical properties of the implants are not the primary focus of this study, the results of the mechanical pull‐out tests (Hung Ta Instrument LTD., Taichung, Taiwan) are summarized in Table S‐1 for reference in supporting material. Notably, Implant II shares the same mechanical performance with Implant I, as it was fabricated by applying a BG coating onto the same base structure as Implant I. The preparation of doped BG is discussed in the following section. Figure 1 provides an overview and scanning electron microscopy (SEM) images (HITACHI S‐3400, Tokyo, Japan) of Implants I, II and III, demonstrating that the printing quality of the BG‐doped Ti‐6Al‐4V powders is acceptable. Figure 1d shows a zoomed‐in SEM image of the surface of Implant II, indicating that the thickness of the BG‐coating is approximately 25 μm.

2.2. Preparation of BGS

The BG powder was synthesized through spray pyrolysis by using a 58S composition (60 mol% silicon dioxide, 35 mol% calcium oxide, and 5 mol% phosphorus pentoxide). The solid precursors included 6.70 g of tetraethyl orthosilicate (99.9 wt%, Showa, Japan), 1.40 g of calcium nitrate tetrahydrate (98.5 wt%, Showa, Japan), and 0.73 g of triethyl phosphate (99.0 wt%, Alfa Aesar, USA). These compounds were mixed with 120.00 g of ethanol containing 3.20 g of 0.5 M diluted hydrochloric acid. The resulting solution was stirred at room temperature for 24 h to ensure homogeneity. The homogeneous solution was then transferred to an ultrasonic atomizer (KT‐100A, King Ultrasonic, New Taipei City, Taiwan) operated at a frequency of 1.67 MHz. The atomized droplets were directed into a tube furnace (D110, Dengyng, New Taipei City, Taiwan) with three heating zones set at 250°C, 550°C, and 300°C for preheating, calcination, and cooling, respectively. At the furnace exit, a high voltage of 16 kV was applied to charge the surface of the powders. The charged powders were subsequently neutralized and condensed within a grounded stainless‐steel electrostatic collector.

2.3. In Vivo Experiments

2.3.1. Animals

All in vivo experiments were conducted in accordance with the ARRIVE guidelines [36]. The procedures for the care and use of research animals adhered to Taiwanese regulations, ISO 10993‐6:2016, and the Good Laboratory Practice for Nonclinical Laboratory Studies (Ministry of Health and Welfare, R.O.C., 3rd ed., 2006). Four male New Zealand rabbits (NZRs) aged between 6 and 7 months (average: 6.5 months) and weighing between 3.2 and 3.6 kg (average: 3.4 kg) were used in this study. NZRs were chosen in the experiments due to their Haversian bone structure similar to that of humans, appropriate body size, ease of handling, and cost‐effectiveness [37]. The femur was selected as the implantation site because of its large bone mass, simple geometry, and clinical relevance. Rabbits underwent surgical procedures and were individually housed in cages at the institute (Master Laboratory Co., LTD, Hsinchu, Taiwan) under controlled environmental conditions, including a temperature of 18°C–21°C, natural lighting, moderate moisture, and appropriate air circulation. During the study period, the rabbits were provided with Prolab Rabbit Diet (Lab Diet, PMI Nutrition International, USA) and were given ad libitum access to water.

2.3.2. Surgical Procedures and Animal Sacrifice

The four NZRs were randomly divided into two groups, and each group was killed at either week 4 or 12 following implantation surgeries. See Table 1, a total of eight implants (four Implant I, two Implant II, and two Implant III) were assigned to the left and right femurs of the rabbits in each group with controlled randomness. They are all implanted in the diaphysis region of the bone. A total of 16 implants were employed (eight for 4 weeks and eight for 12 weeks); ultimately, 32 specimens were obtained (each experimental site has 2 thread regions). After exposing the femurs, an implanter was used to perforate the experimental sites. The implant diameters were precisely matched to the implant beds in the bone cortex to prevent the ingrowth of fibrous tissue. After careful placement of the implants, the surgical wounds were closed, and the rabbits were administered the antibiotic gentamycin (5 mg/kg intramuscularly) for three consecutive days to prevent infection.

Table 1.

Implant distribution in 4 individual animals.

	4 weeks		12 weeks
Animal	A	B	C	D
Right Femur 1	Implant II	Implant I	Implant II	Implant I
Right Femur 2	Implant II	Implant I	Implant II	Implant I
Left Femur 1	Implant III	Implant I	Implant III	Implant I
Left Femur 2	Implant III	Implant I	Implant III	Implant I

Note that, in fact, a priori sample size calculation was performed using G*Power software (large effect size f = 0.4, error probability α = 0.05, power 1‐β = 0.8, number of groups = 3), which suggested a total of 36 samples (12 per group). However, due to research budget constraints, data were collected from a total of 32 thread pairs. Specifically, implant types II and III included 4 implants each, and implant type I included 8 implants. Cross‐sectional images of each implant were analyzed, which captured both the left and right rows of threads, yielding a total of 32 samples, (4 + 4 + 8) × 2, for statistical analysis. This separation between left and right thread regions was maintained during analysis to account for potential asymmetry arising from limb dominance or habitual loading patterns in the animal model.

2.3.3. Histological Processing and Statistical Analysis

To evaluate bone healing in peri‐implant areas, histological analysis was performed at 4 and 12 weeks postimplantation. Specimens retrieved from the femurs were fixed in 10% neutral formalin for 3 days at room temperature. These specimens were then dehydrated in a graded ethanol series (60% to 100%) for 7 days and embedded in polymethyl methacrylate. The embedded blocks were sectioned into 500‐µm‐thick slices by using a low‐speed precision cutter (IsoMet 11‐1280‐170, Buehler, IL, USA). The sections were subsequently ground to a thickness of approximately 5 µm, polished, and stained with aniline blue for optical microscopy.

Two parameters were analyzed, namely, (1) BIC and (2) BD, in the region of interest (ROI), which was defined as the area between the implant threads. Each implant was sectioned at multiple positions, and from these, the two best‐preserved cross‐sections were selected for analysis. Each cross‐sectional image captured both the left and right rows of threads, and analysis was conducted using 40X magnification microscope images as shown in Figure 2a–c, g–i, typically containing approximately five threads per side. As a result, each implant contributed two data sets—one from each side—yielding a total of 64 data points from 16 implants. The ROI is illustrated in Figure 2 as the region between the red dashed line connecting the thread tips and the implant surface. BIC was calculated as the percentage of the total length of the line between the bone (including the newly formed bone, osteoid, and mineralized bone) and the implant relative to the total implant length within the ROI. BD was determined by calculating the percentage of the area occupied by the bone within the ROI. Histomorphometric analysis was conducted using the image postprocessing software ImageJ (U.S. National Institutes of Health, MD, USA).

Figure 2.

Light micrographs (40X) of bone growth around Implants I to III after (a–c) 4 and (g–i)12 weeks; detailed bone/tissue integration within the threads is revealed in the enlarged images (100X), (d–f) and (j–l). Red circles in (d, i) give examples on “high BIC but low BD” phenomenon; dotted red circles in (i) represents “high BD but low BIC” regions. Dashed lines in (g–i) define the ROI for BD and BIC calculations.

A parametric ANOVA was then conducted to investigate the effects of material composition and implant fabrication on bone healing at weeks 4 and 12. First, a global test was performed using the statistical function “f_oneway()” from the SciPy package [38]. Then, pairwise multiple comparisons between groups were conducted using Student's t test from the Scikit‐Posthocs package. Significance was defined as p < 0.05 (p < 0.05*), and p < 0.001 was considered highly significant (p < 0.001**). The correlation between BIC and BD was analyzed using Spearman rank correlation coefficients.

3. Results

3.1. Descriptive Histological Analysis

The healing of the surgical sites progressed uneventfully and without complications in all NZRs. No signs of inflammation were observed around any of the implants. Figure 2 presents light micrographs depicting bone growth around Implants I to III after 4 and 12 weeks. The blue regions in the images represent the bone, and detailed bone integration within the threads is highlighted in the enlarged images. Overall, all three types of implants exhibited substantial bone growth around the middle portions of the implants after 4 weeks of healing. As illustrated in Figure 2b,c, Implants II and III (BG‐coated and BG‐doped Ti‐6Al‐4V) promoted greater bone growth than did Implant I (pure Ti‐6Al‐4V), which led to less new bone tissue in the peri‐implant areas [Figure 2a]. Enlarged images of the threads at the middle portions of the three implants revealed low BIC behaviors (white in the image) near the thread surface in Implant I, which are marked by red circles in Figure 2d. By contrast, improved osseointegration was observed in Implants II and III, as depicted in Figure 2e,f, respectively.

After 12 weeks of healing, a significant volume of new bone formation was observed, particularly around the middle and bottom sections of all three types of implants, as illustrated in Figure 2g–i. However, white regions were present in the newly formed bone areas surrounding Implants II and III (Figure 2h,i). Thus, the average BD values of Implants II and III were lower than that of Implant I. Enlarged images of selected threads (Figure 2j–l) revealed that the surfaces of all three implant types supported effective bone osseointegration.

The numerical histomorphometric results for BD and BIC at weeks 4 and 12 are presented in Table 2. At week 4, Implant II (BG‐coated Ti‐6Al‐4V) had the highest average BD (68.06%) and BIC (67.05%) values, whereas Implant I (pure Ti‐6Al‐4V) had the lowest BD (15.59%) and BIC (13.58%) values. At week 12, the BD and BIC values for Implant II decreased to 44.79% and 49.56%, respectively, whereas those for Implant I significantly increased to 64.3% and 64.07%, respectively. Additionally, Implant III (BG‐doped Ti‐6Al‐4V) exhibited increased values at week 12 (BD: 58.61%; BIC: 48.22%). The reduction in bone volume around Implant II after the fourth week may be attributable to a marginal bone loss effect, which is discussed further in Section 4.

Table 2.

Histomorphometric Rresults of Average BIC and BD Values at Weeks 4 and 12.

Time point	Implant	Average bone density (%)	Average bone in contact (%)
4 week	I	15.59	13.58
	II	68.06	67.05
	III	47.78	40.98
12 weeks	I	64.30	64.07
	II	44.79	49.56
	III	58.61	48.22

3.2. Histomorphometric Analysis of BD and BIC

A parametric ANOVA was conducted to evaluate the effects of the test and control materials at weeks 4 and 12. The analysis began with a global test, followed by post hoc comparisons. The BD and BIC results are presented in Figure 3, where p values smaller than 0.001 are considered highly significant and are denoted by **. BD was significantly associated with the biomaterials and the healing period (p < 0.0001 in ANOVA). In the post hoc tests, at week 4, both Implant II (BG‐coated) and Implant III (BG‐doped) had significantly higher BD values than did Implant I (pure Ti‐6Al‐4V; p < 0.0001 and p < 0.0004, respectively). However, by week 12, the BD values did not significantly differ among the three implant types (p = 1, 0.96, and 0.45, respectively). The BD of Implant I significantly increased between weeks 4 and 12 (p < 0.0001).

Figure 3.

Histomorphometric results for BD and BIC.

BIC exhibited a trend similar to that of BD and was significantly associated with the biomaterials and the healing period (p < 0.0001 in ANOVA). In the post hoc tests, after 4 weeks of healing, Implant II (BG‐coated) had significantly higher BIC values than did Implant I (pure Ti‐6Al‐4V; p < 0.0001). By week 12, the BIC values did not significantly differ among the three implant types (p = 0.65, 0.99, and 0.53, respectively). Consistent with the observations for BD, the BIC value of Implant I significantly increased between weeks 4 and 12 (p < 0.0001).

3.3. Correlation Analysis

A Spearman rank correlation was conducted to evaluate the relationship between BD and BIC at weeks 4 and 12 (Figure 4). At week 4, BD and BIC exhibited strong positive correlations with a high linear dependence (correlation coefficient: +0.96, p < 0.0001). However, at week 12, the correlation weakened to a moderate positive relationship (correlation coefficient: +0.64, p < 0.0001). This decrease is attributable to data points, primarily corresponding to Implant III, that indicated either “high BIC but low BD” or “high BD but low BIC.”

Figure 4.

Correlation between BD and BIC is positive at both week 4 (+0.96) and week 12 (+0.64).

The “high BIC but low BD” scenario indicates the presence of regions where bone adhered well to the implant surface, forming a thin layer, whereas in other peri‐implant areas, limited bone growth was noted. An example of this is indicated by a solid red circle in Figure 2d,i. Notably, “high BD but low BIC” indicated substantial bone growth in peri‐implant areas with minimal contact on the implant surface, which is represented by a dotted red circle in Figure 2i.

4. Discussion

Osseointegration is a bone healing process that reflects the establishment and maintenance of rigid fixation in bone subjected to functional loading [39]. This process occurs in several stages: initial stabilization of the implant under various stimuli, bone regeneration in peri‐implant areas, and structural integration as a cohesive unit [40]. Because the early phase of osseointegration determines primary stability and eventually the success of fixation, the first 12 weeks after implantation are pivotal for evaluating bone ingrowth and remodeling. Although osseointegration is a complex process involving immune‐inflammatory responses, angiogenesis, and osteogenesis [41], cellular activities and bone development can be effectively studied through in vivo experimental observations, particularly by using histological evaluations [42, 43].

In the present histological studies, the limitations of staining techniques and image magnification restricted this study's ability to analyze the quantity and location of osseous apposition with the implants. Empirically, a healing period of 4 to 12 weeks is sufficient to generate substantial amounts of mature lamellar bone and some immature woven bone [31]. Thus, differentiation into fibrous tissue, cartilage, or mature and immature bone types was not classified in this study. Instead, we focused on the volume of bone formation and BIC, which reflect the extent of bone ingrowth. Moreover, the physical and chemical characteristics of the implant surface, including roughness, topography, composition, energy and wettability [44, 45, 46], considerably affect cellular responses and events in peri‐implant areas. Thus, we examined the bone‐implant interface because it directly affects the progress and quality of osseointegration.

Our histological results reveal that Implant II (BG‐coated) demonstrated superior bone regeneration to that of Implants I (pure Ti‐6Al‐4V) and III (BG‐doped) at week 4, as indicated by higher BIC and BV values for Implant II. The ability of BGs to enhance bone formation has been demonstrated in many previous studies [21]. This effect is attributable to the release of ionic products, including apatite, calcium, and silicon ions, from BG [47]. These products can stimulate osteogenic activity by activating biological growth factors associated with bone formation. The dissolution of BG also facilitates the formation of a strong bond between the surrounding bone and the apatite layer on the implant surface. Thus, Implants II and III, both of which contained 58S, promoted rapid mineralization during the early stages of bone regeneration in this study.

In particular, BG‐coated Implant II exhibited better bone ingrowth than BG‐doped Implant III. This difference can be attributed to the larger surface area of BG available in the coating compared with that achievable through the doping method. However, unlike porous scaffolds [48], the implants used in this study were designed without intentional porosity; therefore, no calculation of pore size or porosity was performed. Regarding surface contact, the BG‐doped implants were prepared by mixing submicron‐sized BG particles with Ti‐6Al‐4V powder before printing, as shown in Figure 1e. By examining the SEM image, ~8.71% of the BG‐doped surface is in contact with the surrounding bone. In contrast, the BG‐coated implants exhibit a more uniform surface coverage, as show in Figure (d), and the BG layer is likely to be in full contact with the bone. In addition, the rough surfaces produced by applying AM techniques, such as 3D printing, increase the surface area and enhance interlocking with the living bone [44, 46]. The coated surface further promotes direct contact osteogenesis by supporting greater cellular proliferation and bone integration. However, the effects of BG addition may vary depending on factors such as the coating method, thickness, and composition. This study only reflects BG coating effect at the thickness of approximately 25 μm. Future studies should compare the effects of BG addition through coating and doping methods.

The results of the present study indicate that after 12 weeks of healing, bone ingrowth reached similar levels among the three types of implants. However, pure Ti‐6Al‐4V (Implant I) exhibited higher BIC and BD values than did the two BG‐added implants (Implants II and III). Moreover, numerous bubble‐like white regions were observed in the bone (blue) surrounding Implants II and III, as indicated in Figure 2h,i. These findings suggest that although BG demonstrated the ability to promote osteogenesis and angiogenesis through its high dissolution rate of ions that enhanced biological activities, particularly during the early postoperative stage [49], BG remains an imperfect material for supporting long‐term bone growth. Van Dijk La et al. reported that the fusion rate of 45S5 BGs was inferior to that of autograft after 12 weeks of healing in an ovine posterolateral spinal model [50]. The calcium content in the BGs used in this study may have led to a high pH value. This phenomenon has been reported in the literature [51, 52] and occurs because of the rapid release of calcium, sodium, or other alkaline ions from BGs, leading to an environment that is unfavorable for cellular activity and detrimental to subsequent bone ingrowth. This likely explains why Implant II (BG‐coated) exhibited the lowest average BD value after 12 weeks, whereas the impact on the BD of Implant III (BG‐doped) was less pronounced. To address these challenges, pH‐neutral BGs, as reported in the literature [53, 54, 55], can be used in future applications.

The phenomenon of “high BIC but low BD” and “high BD but low BIC” observed in Implant III suggests that the distribution of BG‐doped particles in the additively manufactured implant is not uniform. This nonuniformity leads to inconsistent bone ingrowth performance around Implant III, as evidenced by the varying bone morphologies observed around its threads in Figure 2i. These bone morphologies include regions with no bone ingrowth (white), a thin layer of bone attached to the implant surface (high BIC but low BD), areas with a thin layer of unattached bone (high BD but low BIC), and threads fully filled with bone. Variations in bone growth around BG‐doped implants could be related to printing or composition defects. This variability highlights the need for further research to improve the quality of additively manufactured BG‐doped implants to achieve more uniform particle distribution and consistent osseointegration performance. For instance, printing parameters (e.g., nozzle temperature, printing speed, layer height) and the development of stable composite formulation can lead to better dispersion of BG particles. In our case, collagen was used as a binder within the metal powder–based composite formulation, suggesting that optimizing binder composition and distribution could contribute to improved homogeneity and printing consistency. Post‐processing treatments are also suggested to enhance the quality [56].

This study revealed the effect of BG addition during the early stages of bone healing. However, further in‐depth investigations are required. For example, the present study used only four rabbits in its animal tests; a larger sample size is required to achieve statistical significance and ensure generalizability for clinical applications. In addition, employing more advanced staining techniques in future studies could provide valuable insights into bone maturity and cell differentiation. Future experiments could also include pH value measurements to optimize the composition of BGs. For additively manufactured implants, printing parameters are crucial because they affect surface conditions such as roughness and wettability. Thus, in addition to material composition, different surface conditions should be considered. In terms of implant mechanical testing, although push‐out testing was conducted in this study to assess the strength of the implant and the corresponding fixation, torque testing may provide additional insights for threaded designs. Incorporating torque‐based evaluations in future studies may offer a more comprehensive understanding of implant stability in clinical settings. Finally, extending the healing period in future studies would be advantageous for understanding the long‐term remodeling process. According to previous studies on osseointegration with implants in rabbits [51, 52], studies spanning 12 to 24 weeks could provide comprehensive insights into the progression of bone healing and integration.

5. Conclusion

Compared with implants made of pure Ti‐6Al‐4V, 3D‐printed implants made of Ti‐6Al‐4V with 58S BGs added through either coating or doping are associated with enhanced bone ingrowth during the early stages of bone healing, as evidenced by the in vivo BD and BIC results. This finding is notable because rapid osseointegration is highly desirable in clinical applications. Furthermore, the current study demonstrated that BG can be effectively doped onto conventional 3D printing particles, such as Ti‐6Al‐4V, and that these doped particles can be successfully utilized in AM. Although BG‐doped implants accelerate early‐stage bone healing, their bone ingrowth performance remains inconsistent over the long term because of the nonuniform distribution of BG within the implant. Addressing challenges such as optimizing the pH balance of BG and achieving a consistent doping concentration is critical. If these problems are resolved, the proposed biomaterial and its applications hold considerable potential for becoming viable commercial implant materials in the future.

6. Social Media Promotion

This study investigates the effects of bioactive glass (BG) incorporation methods on osseointegration in 3D‐printed Ti‐6Al‐4V implants. Using selective laser melting, we compared BG doping and BG coating, and evaluated in vivo performance over 4 and 12 weeks. Results show that both approaches enhance early‐stage bone integration, with BG coating showing greater initial improvement.

Author Contributions

Chih‐Yu Lee wrote the initial manuscript, prepared figures, and collected and analyzed the data. Pei‐Ching Kung analyzed the data. Chih‐Chieh Huang collected the data, established the model, and executed the majority of the experiments. San‐Yuan Chen, Shao‐Ju Shih, E‐Wen Huang, and Meng‐Huang Wu established the model and offered the concept and resources. Meng‐Huang Wu executed the majority of the experiments, and supervised the study. Nien‐Ti Tsou wrote the initial manuscript, established the model, supervised the study, and made a contribution to the revision of the manuscript. All authors have read and approved the final submitted manuscript.

Supporting information

Supplementary Material Table S1: Mechanical pull‐out test. Noted that no data is measured with Implant II as we assumed BG coating doesn't affect the bulk mechanical properties.