Cell–cell interaction between vocal fold fibroblasts and bone marrow mesenchymal stromal cells in three‐dimensional hyaluronan hydrogel

Mesenchymal stromal cells (MSCs) are multipotential adult cells present in all tissues. Paracrine effects and differentiating ability make MSCs an ideal cell source for tissue regeneration. However, little is known about how interactions between implanted MSCs and native cells influence cellular growth, proliferation, and behaviour. By using an in vitro three‐dimensional (3D) co‐culture assay of normal or scarred human vocal fold fibroblasts (VFFs) and bone marrow‐derived MSCs (BM‐MSCs) in a uniquely suited hyaluronan hydrogel (HyStem–VF), we investigated cell morphology, survival rate, proliferation and protein and gene expression of VFFs and BM‐MSCs. BM‐MSCs inhibited cell proliferation of both normal and scarred VFFs without changes in VFF morphology or viability. BM‐MSCs demonstrated decreased proliferation and survival rate after 7 days of co‐culture with VFFs. Interactions between BM‐MSCs and VFFs led to a significant increase in protein secretion of collagen I and hepatocyte growth factor (HGF) and a decrease of vascular endothelial growth factor (VEGF), monocyte chemotactic protein‐1 (MCP‐1) and interleukin‐6 (IL‐6). In particular, BM‐MSCs significantly upregulated matrix metalloproteinase 1 (MMP1) and HGF gene expression for scarred VFFs compared to normal VFFs, indicating the potential for increases in extracellular matrix remodelling and tissue regeneration. Application of BM‐MSCs‐hydrogels may play a significant role in tissue regeneration, providing a therapeutic approach for vocal fold scarring. Copyright © 2013 John Wiley & Sons, Ltd.     

Ultrasonic recording of the fundamental frequency of a voice during normal speech

A continuous wave of ultrasound through larynx is used to determine the fundamental frequency of the vocal folds during normal speech. The variation of the ultrasound intensity transmitted between two matched transducers through the vocal fold level is recorded. The apparatus used for these recordings is described. Different transducer positions are investigated, and further, the transducer geometry is discussed.     

The efficacy of a novel collagen–gelatin scaffold with basic fibroblast growth factor for the treatment of vocal fold scar

Vocal fold scar remains a therapeutic challenge. Basic fibroblast growth factor (bFGF) was reported to have regenerative effects for vocal fold scar, although it has the disadvantage of rapid absorption in vivo. A collagen–gelatin sponge (CGS) can compensate for the disadvantage by providing a sustained release system. The current study evaluated the efficacy of CGS combined with bFGF on vocal fold scar, using rat fibroblasts for an in vitro model and a canine in vivo model. We prepared fibroblasts from scarred vocal folds (sVFs) in rats and showed that bFGF accelerated cell proliferation and suppressed expression levels of cleaved caspase 3 and α‐smooth muscle actin. Has 1, Has 3, Fgf2, Hgf and Vegfa mRNA levels were significantly upregulated, while Col1a1 and Col3a1 were dose‐dependently downregulated, with a maximum effect at 100 ng/ml bFGF. In an in vivo assay, 6 weeks after lamina propria stripping, beagles were divided into three groups: CGS alone (CGS group); CGS with bFGF (7 µg/cm²; CGS + bFGF group); or a sham‐treated group. Vibratory examination revealed that the glottal gap was significantly reduced in the bFGF group and the two implanted groups, whereas the CGS + bFGF group showed higher mucosal wave amplitude. Histological examination revealed significantly restored hyaluronic acid and elastin redistribution in the CGS + bFGF group and reductions in dense collagen deposition. These results provide evidence that CGS and bFGF combination therapy may have therapeutic potential and could be a promising tool for treating vocal fold scar. Copyright © 2015 John Wiley & Sons, Ltd.     

Tissue engineering of the larynx: A contemporary review

ObjectiveTissue engineering has been a topic of extensive research in recent years and has been applied to the regeneration and restoration of many organs including the larynx. Currently, research investigating tissue engineering of the larynx is either ongoing or in the preclinical trial stage.MethodsA literature search was performed on the Advanced search field of PubMed using the keywords: “(laryncheal tissue engineering) AND (cartilage regeneration OR scaffolds OR stem cells OR biomolecules).” After applying the selection criteria, 65 articles were included in the study.ResultsThe present review focuses on the rapidly expanding field of tissue‐engineered larynx, which aims to provide stem cell–based scaffolds combined with biological active factors such as growth factors for larynx reconstruction and regeneration. The trend in recent studies is to use new techniques for scaffold construction, such as 3D printing, are developed. All of these strategies have been instrumental in guiding optimization of the tissue‐engineered larynx, leading to a level of clinical induction beyond the in vivo animal experimental phase.ConclusionsThis review summarizes the current progress and outlines the necessary basic components of regenerative laryngeal medicine in preclinical fields. Finally, it considers the design of scaffolds, support of growth factors, and cell therapies toward potential clinical application.     

喉部肿块螺旋CT三维成像的应用

目的　探讨螺旋CT三维成像表面覆盖 (shadedsurfacedisplay ,SSD)技术在喉部肿块的初步临床应用。 方法　3 1例全部采用螺旋CT扫描技术 ,确定扫描层面与声带水平 ,范围从会厌软骨至环状软骨 ,正常喉的SSD重建所见与喉镜所见进行比较 ,喉部肿块的SSD重建所见与喉镜、手术、病理所见进行对照分析。结果　SSD重建技术能清楚显示正常喉及下咽部的腔内结构 ,通过增减重建范围、削割、旋转、Light等软件功能的应用 ,有利于喉腔内部结构、病变部位、形态等解剖关系的显示。结论　SSD重建技术是普及型螺旋CT在不具备工作站与仿真内镜软件下就能完成的一项技术 ,能显示正常和异常喉及下咽部的腔内结构 ,具有仿真内镜类似的效果 ,是一种喉非创伤性、操作简单的新技术。     

In vivo implantation of a tissue engineered stem cell seeded hemi‐laryngeal replacement maintains airway,phonation, and swallowing in pigs

Laryngeal functional impairment relating to swallowing, vocalisation, and respiration can be life changing and devastating for patients. A tissue engineering approach to regenerating vocal folds would represent a significant advantage over current clinical practice. Porcine hemi‐larynx were de‐cellularised under negative pressure. The resultant acellular scaffold was seeded with human bone marrow derived mesenchymal stem cells and primary human epithelial cells. Seeded scaffolds were implanted orthotopically into a defect created in the thyroid cartilage in 8 pigs and monitored in vivo for 2 months. In vivo assessments consisted of mucosal brushing and bronchoscopy at 1, 2, 4, and 8 weeks post implantation followed by histological evaluation post termination. The implanted graft had no adverse effect on respiratory function in 6 of the 8 pigs; none of the pigs had problems with swallowing or vocalisation. Six out of the 8 animals survived to the planned termination date; 2 animals were terminated due to mild stenosis and deep tissue abscess formation, respectively. Human epithelial cells from mucosal brushings could only be identified at Weeks 1 and 4. The explanted tissue showed complete epithelialisation of the mucosal surface and the development of rudimentary vocal folds. However, there was no evidence of cartilage remodelling at the relatively early censor point. Single stage partial laryngeal replacement is a safe surgical procedure. Replacement with a tissue engineered laryngeal graft as a single procedure is surgically feasible and results in appropriate mucosal coverage and rudimentary vocal fold development.     

In Vitro Ultrasound Biomicroscopic Imaging of Colitis in Rats

Objective. The purpose of this study was to show the feasibility of 50‐MHz ultrasound biomicroscopy (UBM) to image the rat colon. Methods. B‐mode images were obtained from ex vivo colon samples (n = 4) collected from Rattus norvegicus (Berkenhout, 1769) rats, with 2,4,6‐trinitrobenzene sulfonic acid–induced colitis in 3 of them. Left colon rectangular fragments (5 × 5 mm) were obtained after necropsy, and UBM images were acquired with the samples immersed in saline at 37°C. All layers of the normal intestinal wall were analyzed according to their thickness and the presence of uneven bowel mucosa (ulcers). The folds and layers detected by UBM were correlated with histopathologic analysis. Results. The 4 layers of the normal colon were identified on the UBM images: the mucosa (hyperechoic), muscularis mucosae (hypoechoic), submucosa (hyperechoic), and muscularis externa (hypoechoic). On 2 UBM images, superficial ulcers were detected, approximately 0.5 mm in size, with intestinal involvement limited to the mucosa. The histopathologic analysis verified enlargement of submucosa layers due to an edema associated with sub‐mucosa leukocyte infiltration. On 1 UBM image, it was possible to detect a deep ulcer, which was confirmed by the light microscopic analysis. Conclusions. An ultrasound imaging system was scaled and optimized to visualize the rat colon. Ultrasound biomicroscopy provided axial and lateral resolutions close to 25 and 45 μm, respectively, and adequate penetration depth to visualize the whole thickness of an inflamed colon. The system identified the colon layers and was able to detect mural changes and superficial ulcers on the order of 500 μm.     

A phase I/II exploratory clinical trial for intracordal injection of recombinant hepatocyte growth factor for vocal fold scar and sulcus

Vocal fold scar and sulcus are intractable diseases with no effective established treatments. Hepatocyte growth factor (HGF) has preclinically proven to have potent antifibrotic and regenerative effects on vocal fold scar. The current Phase I/II clinical trial aims to examine the safety and effectiveness of intracordal injection of a recombinant human HGF drug for patients with vocal fold scar or sulcus. This is an open‐label, dose‐escalating, first‐in‐human clinical trial. Eighteen patients with bilateral vocal fold scar or sulcus were enrolled and divided into three groups: Step I received 1 μg of HGF per vocal fold; Step II received 3 μg of HGF; and Step III received 10 μg of HGF. Injections were administered once weekly for 4 weeks. The protocol treatment was performed starting with Step I and escalating to Step III. Patients were followed for 6 months post‐treatment. Local and systemic safety aspects were examined as primary endpoints, and therapeutic effects were assessed as secondary endpoints using voice handicap index‐10; maximum phonation time; vocal fold vibratory amplitude; grade, rough, breathy, asthenic, strained scale; and jitter. The results indicated no serious drug‐related adverse events in either the systemic or local examinations. In whole‐subject analysis, voice handicap index‐10, vocal fold vibratory amplitude, and grade, rough, breathy, asthenic, strained scale were significantly improved at 6 months, whereas maximum phonation time and jitter varied. There were no significant differences in phonatory data between the step groups. In conclusion, intracordal injection of a recombinant human HGF drug was safe, feasible, and potentially effective for human patients with vocal fold scar or sulcus.     

科学发声矫治法在显微喉镜下声带息肉切除术后康复中的作用

目的探讨科学发声矫治法在声带息肉患者术后康复中的作用。方法选取60例声带息肉术后患者,随机分为A、B两组,A组为综合治疗组,B组为综合治疗＋科学发声矫治法组,通过术后1、3、6个月门诊进行复查,观察患者术后的恢复情况。结果科学发声矫治法B组在音质改善及喉镜下声带黏膜的恢复情况明显好于A组（P〈0.05）。结论科学发声矫治法可以改善声带息肉术后患者的发音质量并降低其复发率。     

Noninvasive assessment of laryngeal phonation function using color Doppler ultrasound imaging

Color Doppler imaging (CDI) was used to identify the morphology of vocal folds (VF) and to quantify the tissue horizontal displacement velocity (HDV) in the vibrating portion of VF. Mucosal HDV that gives an estimate of the stiffness of the VF cover is very important clinically. The VF and its cover were shown to be very hypoechoic and not adequately visible in B-scan image. However, in this study, we found that the structure of the true VF, especially the mucosa and the superficial layer of the lamina propria, the so-called "cover," can be easily identified and evaluated using CDI. The mean VF displacement velocity was measured by decoding the pseudocolor codes of the ultrasound (US) image at the vibrating sites. The mucosal mean HDV obtained from 10 normal men of age 34 +/- 8 years phonating at their most comfortable pitch and intensity (98 +/- 12 Hz, 55-65 dB) was about 68 +/- 10 cm/s, which agreed reasonably with the literature. Therefore, the CDI could be used as a potential tool for assessing the phonation function in the larynx nonintrusively.     

Features of in vitro ultrasound biomicroscopic imaging and colonoscopy for detection of colon tumor in mice

The present work tested the capability of ultrasound biomicroscopy (UBM), at 45 MHz, to provide cross-sectional images with appropriate resolution and contrast to detect tumors and determine their penetration depths on the colon of mice, Mus musculus (Linnaeus 1758), treated with carcinogen for colon tumor induction. B-mode images were obtained, in vitro, from each animal (13 treated and 4 untreated) colon opened longitudinally and immersed in saline solution at room temperature. Prior to UBM inspection, all animals were also examined by colonoscopy. The layers of normal colon identified by UBM are: mucosa (hyperechoic), muscularis mucosae (hypoechoic), submucosa (hyperechoic) and muscularis externa (hypoechoic). UBM images of colon lesions presented structures corresponding to tumors (hyperechoic), lymphoid hyperplasia (hypoechoic) and polypoid tumors (hyperechoic). Additionally, tumoral lesion invasion through the colon was also identified. When compared with histopathologic analysis, all colon lesions detected by UBM were confirmed, while colonoscopic findings had two false negatives.     

Vibration stimulates vocal mucosa‐like matrix expression by hydrogel‐encapsulated fibroblasts

The composition and organization of the vocal fold extracellular matrix (ECM) provide the viscoelastic mechanical properties that are required to sustain high‐frequency vibration during voice production. Although vocal injury and pathology are known to produce alterations in matrix physiology, the mechanisms responsible for the development and maintenance of vocal fold ECM are poorly understood. The objective of this study was to investigate the effect of physiologically relevant vibratory stimulation on ECM gene expression and synthesis by fibroblasts encapsulated within hyaluronic acid hydrogels that approximate the viscoelastic properties of vocal mucosa. Relative to static controls, samples exposed to vibration exhibited significant increases in mRNA expression levels of HA synthase 2, decorin, fibromodulin and MMP‐1, while collagen and elastin expression were relatively unchanged. Expression levels exhibited a temporal response, with maximum increases observed after 3 and 5 days of vibratory stimulation and significant downregulation observed at 10 days. Quantitative assays of matrix accumulation confirmed significant increases in sulphated glycosaminoglycans and significant decreases in collagen after 5 and 10 days of vibratory culture, relative to static controls. Cellular remodelling and hydrogel viscosity were affected by vibratory stimulation and were influenced by varying the encapsulated cell density. These results indicate that vibration is a critical epigenetic factor regulating vocal fold ECM and suggest that rapid restoration of the phonatory microenvironment may provide a basis for reducing vocal scarring, restoring native matrix composition and improving vocal quality. Copyright © 2009 John Wiley & Sons, Ltd.     

喉内窥镜手术治疗声带病变的临床研究

目的：探讨喉内窥镜手术治疗声带病变的效果。方法：对120例声带病变患者采用无插管全麻支撑喉镜下喉内镜手术治疗,并对其临床资料进行分析。结果：120例近期（1个月内）疗效为：治愈82例,占68．3％,好转32例,占26．7％;无效6例,占5％。远期（1年以）疗效为：治愈９５％例,占７９．２％;好转１６例,占１３．３％,无效９例,占７．５％。结论：无插管全麻支撑喉镜下喉内窥镜治疗声带病变,其中对声带息肉、声带小结效果较好;对喉Ｒeinke's水肿及声带囊肿也有一定的效果;对喉癌及喉乳头状瘤采用内窥镜进行病变范围的观察及活检术是有价值的,但所进行微波组织凝固术治疗喉癌或乳头状瘤只是一种试尝、其疗效尚需进一步观察。     

正常人胸声区不同音频发音时电声门图参数变化

背景国内利用电声门图对正常人喉嗓音进行生理研究的报道较少.目的通过电声门图检测,了解不同年龄、性别健康人在胸声区以不同音频发音所产生的声带振动变化规律.进一步拓展喉嗓音的探索性研究.设计相互对照的开放实验.地点和对象实验在重庆市第三人民医院耳鼻咽喉-头颈外科完成,对象为重庆地区健康受试者120例,其中老年组60例,男女各30例,年龄60～70岁;成年组60例,男女各30例,年龄20～40岁.干预利用电声门图对120例正常人在胸声区以不同音频发音进行检测.主要观察指标声带的接触率、接触幂、接触率微扰、接触幂微扰.结果全部受试者在胸声区随音频由低到高发音,声带接触率逐渐下降,差异具有显著性意义;老年组与成年组比较、男性组与女性组比较,各音频段接触率差异亦均有显著性意义;各组接触幂、接触率微扰、接触幂微扰则差异无显著性意义.结论不同年龄、性别的健康人在胸声区随音频由低到高发音,声带的闭合程度不同,而声带对称性、声带振动规律性在不同性别、年龄的健康人中则并无差异.提示通过电声门图检测对喉嗓音生理研究具有一定意义.     

Paraglottic space schwannoma: a case report and literature review

Neurogenic tumors located in the larynx are extremely rare. Among them, schwannoma is a benign encapsulated tumor originating from Schwann cells, which form nerve fiber sheaths in the peripheral nervous system. We herein report a case of a schwannoma arising from a rare subsite of the larynx and review the literature on laryngeal schwannoma. The case involved a woman with a 1-month history of globus pharyngeus and dysphagia without dysphonia. Rigid laryngoscopy and magnetic resonance imaging showed a large submucosal bulge toward the medial wall of the right pyriform fossa, pushing the right false and true vocal cords and aryepiglottic fold inward. A transcervical approach was used to completely excise the tumor without incisional biopsy or preliminary tracheotomy. Histology confirmed a benign schwannoma originating from the right paraglottic space, which was extremely rare. During follow-up, no evidence of recurrence or a residual mass was found. The transcervical approach is a useful and less invasive treatment for laryngeal schwannoma located in the paraglottic space.     

Two-stage approach for class II mandibular furcation defect with insufficient keratinized mucosa: a case report with 3 years’ follow-up

Periodontal regenerative treatment is useful for intrabony defects and furcation involvement, but is difficult when there is insufficient keratinized mucosa to cover and maintain the regenerative material, particularly in the mandibular molar region. We report the case of a 27-year-old woman who underwent a two-stage surgical approach for a class II furcation defect with gingival recession and insufficient keratinized mucosal width (KMW) and vestibular depth at the mandibular left first molar. We first improved the KMW and keratinized mucosal thickness using an epithelial embossed connective tissue graft with enamel matrix derivative, and then focused on periodontal regeneration at the furcation defect using an enamel matrix derivative and a bovine-derived xenograft. Probing depth reduction, clinical attachment gain, horizontal probing depth reduction, KMW gain, and gingival recession reduction were observed 3 years postoperatively. This case report suggests that this novel staged approach may be effective for treating furcation defects with insufficient keratinized mucosa, thus providing useful insights into periodontal regeneration therapy.     

Dermal matrix scaffold engineered with adult mesenchymal stem cells and platelet-rich plasma as a potential tool for tissue repair and regeneration

The purpose of this study was to investigate the efficacy of Integra, an artificial dermal matrix used as a dermal template for skin regeneration, to form a multifunctional scaffold with human bone marrow-derived mesenchymal stem cells (hMSCs) and platelet-rich plasma (PRP) for tissue engineering and regenerative technology. First, we showed that PRP, used as a supplement for growth medium, represented an optimal substitute for animal serum as well as a source of multiple growth factors, was able to satisfactorily support cell viability and cell proliferation and influence stemness gene expression in hMSCs. Moreover, Integra appeared to be a suitable substrate for hMSCs colonization, as judged by two-photon microscopy combined with fluorescence lifetime imaging (FLIM) and confocal analysis. The cells were then seeded on Integra + PRP for 24 and 48 h. Notably, in these conditions, the seeded cells exhibited a greater aptitude to colonize the scaffold, showed improved cell adhesion and spreading as compared with those cultured on Integra alone, and acquired a fibroblast-like phenotype, indicating that the bioengineered scaffold provided an appropriate environment for cellular growth and differentiation. In conclusion, these results, although preliminary, provide clues for the design of new therapeutic strategies for skin regeneration, consisting in the combination of mesenchymal stem cells with engineered biomaterials.     

Bone regeneration in minipigs via calcium phosphate cement scaffold delivering autologous bone marrow mesenchymal stem cells and platelet‐rich plasma

Macroporous calcium phosphate cement (CPC) with stem cell seeding is promising for bone regeneration. The objective of this study was to investigate the effects of co‐delivering autologous bone marrow mesenchymal stem cells (BMSCs) and autologous platelet‐rich plasma (PRP) in CPC scaffold for bone regeneration in minipigs for the first time. Twelve female adult Tibet minipigs (12–18 months old) were used. A cylindrical defect with 10 mm height and 8 mm diameter was prepared at the femoral condyle. Two bone defects were created in each minipig, one at each side of the femoral condyle. Three constructs were tested: (1) CPC scaffold (CPC control); (2) CPC seeded with BMSCs (CPC‐BMSC); (3) CPC seeded with BMSCs and PRP (CPC‐BMSC‐PRP). Two time points were tested: 6 and 12 weeks (n = 4). Good integration of implant with surrounding tissues was observed in all groups. At 12 weeks, the CPC‐BMSC‐PRP group had significantly less residual CPC remaining in the defect than the CPC‐BMSC group and the CPC control (p < 0.05). The residual CPC volume for the CPC‐BMSC‐PRP group was half that of the CPC control. New bone formation for CPC‐BMSC‐PRP was more than two‐fold that of the CPC control (p < 0.05). CPC‐BMSC‐PRP had new blood vessel density that was nearly two‐fold that of the CPC control (p < 0.05). In conclusion, CPC scaffold with autologous BMSC‐PRP doubled the new bone regeneration and blood vessel density in minipigs compared with the CPC control. In the present study, the new macroporous CPC system with co‐delivered BMSC‐PRP has been shown to promote scaffold resorption and bone regeneration in large defects. Copyright © 2017 John Wiley & Sons, Ltd.     

Repair of surgically created diaphragmatic defect in rat with use of a crosslinked porous collagen scaffold

Large defects in congenital diaphragmatic hernia are closed by patch repair, which is associated with a high complication risk and reherniation rate. New treatment modalities are warranted. We evaluated the feasibility of using an acellular biodegradable collagen bioscaffold for a regenerative medicine approach to close a surgically created diaphragmatic defect in a rat model. Scaffold degradation, cellular ingrowth and regeneration of the diaphragm were studied. In 25 rats, a subcostal incision was made and one third of the right hemidiaphragm was resected. Crosslinked porous type I collagen scaffolds (Ø ~ 14 mm) were sutured into the lesion. Rats were sacrificed at 2, 4, 8, 12 or 24 weeks after scaffold implantation. Implants were evaluated macroscopically and (immuno)histologically. Survival after surgery was 88% with no evidence of reherniation. Histological examination showed that the collagen scaffold degraded slowly and new collagen, elastin and mesothelium were deposited. Blood vessels were observed primarily at the outer borders of the scaffold; their number gradually increased in time. Muscle fibres were found on the scaffold covering up to 10% of the defect. Macroscopically, adhesion of the scaffold to the liver was observed. Use of a collagen scaffold to close a surgically created diaphragmatic defect is feasible, with evidence of new tissue formation. The use of crosslinked collagen scaffolds allows targeted modification; e.g. addition of growth factors to further stimulate growth of muscle cells. Copyright © 2012 John Wiley & Sons, Ltd.     

Rapid healing of a critical‐sized bone defect using a collagen‐hydroxyapatite scaffold to facilitate low dose,combinatorial growth factor delivery

The healing of large, critically sized bone defects remains an unmet clinical need in modern orthopaedic medicine. The tissue engineering field is increasingly using biomaterial scaffolds as 3D templates to guide the regenerative process, which can be further augmented via the incorporation of recombinant growth factors. Typically, this necessitates supraphysiological doses of growth factor to facilitate an adequate therapeutic response. Herein, we describe a cell‐free, biomaterial implant which is functionalised with a low dose, combinatorial growth factor therapy that is capable of rapidly regenerating vascularised bone tissue within a critical‐sized rodent calvarial defect. Specifically, we demonstrate that the dual delivery of the growth factors bone morphogenetic protein‐2 (osteogenic) and vascular endothelial growth factor (angiogenic) at a low dose (5 μg/scaffold) on an osteoconductive collagen‐hydroxyapatite scaffold is highly effective in healing these critical‐sized bone defects. The high affinity between the hydroxyapatite component of this biomimetic scaffold and the growth factors functions to sequester them locally at the defect site. Using this growth factor‐loaded scaffold, we show complete bridging of a critical‐sized calvarial defect in all specimens at a very early time point of 4 weeks, with a 28‐fold increase in new bone volume and seven‐fold increase in new bone area compared with a growth factor‐free scaffold. Overall, this study demonstrates that a collagen‐hydroxyapatite scaffold can be used to locally harness the synergistic relationship between osteogenic and angiogenic growth factors to rapidly regenerate bone tissue without the need for more complex controlled delivery vehicles or high total growth factor doses.