Inhibition of procollagen C-proteinase reduces scar hypertrophy in a rabbit model of cutaneous scarring

Hypertrophic scarring, which results from excessive collagen deposition at sites of dermal wound repair, can be functionally and cosmetically debilitating to the surgical patient. Pharmacological regulation of collagen synthesis and deposition is a direct approach to the control of scar tissue formation. One of the key steps in collagen stabilization is the cleavage of the C-terminal propeptide from the precursor molecule to form collagen fibrils, a reaction catalyzed by procollagen C-proteinase (PCP). We tested the ability of a PCP inhibitor to reduce hypertrophic scar formation in a rabbit ear model. After the placement of four, 7-mm dermal wounds on each ear, New Zealand white rabbits received PCP inhibitor subcutaneously in the left ear at four time points postwounding: days 7, 9, 11, 13 (early treatment; n=20 wounds) or days 11, 13, 15, 17 (late treatment; n=20 wounds). The right ear of each animal served as a control (vehicle alone). Wounds were harvested on postoperative day 28 and scar hypertrophy quantified by measurement of the scar elevation index. Early treatment of wounds with PCP inhibitor did not reduce scar formation compared with controls (p>0.05). However, late treatment resulted in a statistically significant reduction in the scar elevation index (p<0.01). Our results point not only to the potential use of PCP inhibitors to mitigate hypertrophic scarring but also to the temporal importance of drug delivery for antiscarring therapy.     

Inhibition of hypertrophic scar formation with oral asiaticoside treatment in a rabbit ear scar model

Hypertrophic scar (HS) is a fibrotic skin disease characterised by over‐productive collagen and excessive inflammatory reaction, which can be functionally and cosmetically problematic. A scar‐prone constitute will accelerate HS formation and functional disorder, which deserves systemic therapy with oral medicine. To examine the oral therapeutic effectiveness on HS with convincing evidence of gross view and histological improvement, a rabbit ear HS model was employed with oral administration of asiaticoside (AS) at the doses of 12 and 24 mg kg⁻¹ d⁻¹ daily for 60 consecutive days. Gross observation and histological findings showed that oral AS treatment could significantly inhibit HS formation in a dose dependent manner. Semi‐quantification of scar elevation index at days 7, 15, 30, and 60, and quantitative polymerase chain reaction at days 30 and 60 also provided the evidences of reduced scar thickness and inhibited fibrotic gene expressions of collagens I, III, TGF‐β1, interleukins 1β, 6 and 8, and enhanced gene expression of SMAD 7 and PPAR‐γ with a dose‐dependent manner. These results indicated that AS is likely to serve as a systemic therapeutic agent of HS treatment for those who may have scar‐prone constitute via anti‐inflammation, inhibiting fibrotic process, and enhancing matrix degradation.     

他克莫司抑制兔耳瘢痕增生的作用及其机制研究

目的：他克莫司是临床广泛应用的一种药物,其对增生性瘢痕是否产生作用并无相关报道,为此提出并证实了他克莫司可以抑制兔耳瘢痕增生。方法：建立兔耳增生性瘢痕模型,选10只新西兰大耳白兔双耳腹侧用打孔器制作直径1cm圆形创面,伤后14天创面上皮化后给药,每只兔左耳为空白对照组涂等剂量凡士林软膏,右耳为他克莫司治疗组。分别在伤后14天、21天2、8天3、5天和49天采集标本,行HE染色,观察形态学差异;Real-t i me PCR检测纤维化相关因子TGF-β1、TGF-β2、Col l agen-α1等的表达。结果：HE染色可见他克莫司组成纤维细胞数量及胶原纤维较对照组明显减少,PCR结果可见TGF-β1、TGF-β2及Col l agen-α1表达较对照组在各时间点均减少。结论：实验组较对照组瘢痕明显减轻,证明他克莫司显著抑制兔耳瘢痕增生,可作为临床上治疗及预防瘢痕增生的全新疗法。     

Inhibition of pseudophakic endophthalmitis in a rabbit model

A clear lens extraction with insertion of a posterior chamber intraocular lens (PC-IOL) was performed in one eye of each of 20 rabbits. An intracameral injection of 50,000 organisms of gentamicin-sensitive Staphylococcus aureus was given before completion of the surgical procedure. At the completion of surgery, a subconjunctival injection of either gentamicin (20 mg) or gentamicin vehicle was administered randomly, in a masked fashion, to the operative eye. Rabbits were observed daily for signs of endophthalmitis. Vitreous taps were performed on all operative eyes. Ninety percent (p = .001) of control eyes became clinically infected and 80% (p = .007) of control eyes grew S. aureus from their vitreous aspirate. None of the gentamicin-treated eyes became clinically or microbiologically infected with S. aureus.     

The effect of surgical denervation on prevention of excessive dermal scarring: A study on rabbit ear hypertrophic scar model

Background

Previous reports have suggested that the extent of wound contraction, epithelisation and total healing time were influenced by denervation of tissues. In this article, we studied for the first time the effect of sensory denervation on prevention of excessive dermal scarring.

Materials and Methods

Sixteen New Zealand white rabbits were used. Denervation of the right ears was performed by surgical excision of two main sensory nerves. Dissections were also performed on left ears without any nerve excision for the control group. After 14 days of follow-up and confirmation of tissue denervation, an excessive dermal scarring model as defined by Morris et al. was made by surgery on both ears. Twenty-eight days after making the wounds, the tissues were extirpated for analyses. The scars were evaluated by the scar elevation index (SEI), epithelisation time and inflammatory cell count.

Results

The SEI of the denervated side scars was significantly lower than that of the non-denervated side. The rate and timing of total epithelisation and inflammatory cell count between groups yielded no difference.

Conclusions

In this study, the surgical denervation skin reduced scarring. It was suggested that understanding the exact role of sensory nerves and neural mediators in excessive dermal scarring is necessary for the prevention and treatment of scarring.     

Adenovirus-mediated METH1 gene expression inhibits hypertrophic scarring in a rabbit ear model

Hypertrophic scarring remains a major problem for patients who have suffered from surgeries or burns. Vascularization plays an important role in the early phase of hypertrophic scarring. Therefore, the inhibition of angiogenesis might be used as a preventive strategy. In this study, we assessed the effect of anti-angiogenesis resulting from adenovirus-mediated METH1 (metalloprotease and thrombospondin1) gene expression on the hypertrophic scar formation in a rabbit ear model of hypertrophic scarring. We first investigated the number of microvessel and microcirculatory perfusion in untreated scars on days 10, 30, 60, and 90 after epithelialization. Then, we examined the effect of anti-angiogenesis by adenovirus-mediated METH1 expression on hypertrophic scar formation by calculating the scar elevation index, counting the microvessel and argyrophilic nucleolar organizer region particle, and detecting the amount of collagen on days 30 and 60 after treatment. We found that untreated scar tissues at the proliferative phase (days 10–60 after epithelialization) had a significantly higher density of microvessel and microcirculatory perfusion than those at the mature phase (day 90 after epithelization) (both p <0.05). On days 30 and 60 after treatment, the hypertrophic scar formation was significantly inhibited in the treatment group. There was significantly reduced scar elevation index, microvessel count, number of argyrophilic nucleolar organizer region, and total collagen content for treated scars. Our results demonstrate that METH1 has a markedly inhibitive effect on the formation of hypertrophic scar, and may thus have a promising application in the prevention of human hyperthropic scars.     

光动力学疗法对兔耳增生性瘢痕作用的初步研究

目的 探讨光动力学疗法对兔耳增生性瘢痕的作用。方法 建立兔耳增生性瘢痕模型后，将增生性瘢痕块随机分为血卟啉单甲醚（HMME）一光动力学疗法（PDT）治疗组和对照组（n=12）。观察瘢痕生长情况，建模术后28d取材行常规HE染色和胶原纤维VG染色，计算微血管密度，观察PDT对瘢痕的影响。结果 PDT治疗后瘢痕厚度显著降低，真皮层变薄；微血管数量及成纤维细胞数量减少，且主要位于真皮深层；胶原含量下降，胶原纤维排列较规则有序。结论 在瘢痕形成早期，HMME—PDT能够有效抑制兔耳瘢痕的增生，该方法有可能成为瘢痕预防与治疗的有效方法。     

Surgical capsular release reduces flexion contracture in a rabbit model of arthrofibrosis

Animal models of joint contracture may be used to elucidate the mechanisms of arthrofibrosis. Patients with joint contracture commonly undergo surgical capsular release. Previous animal models of joint contracture do not simulate this aspect of arthrofibrosis. We hypothesize that a surgical capsular release will decrease the severity of arthrofibrosis in this rabbit model. A capsular contracture was surgically created in 20 skeletally mature rabbits. Eight weeks later, ten rabbits underwent capsular release, which consisted of elevation of the posterior capsule through a lateral incision and manipulation under anesthesia. Ten rabbits had a sham incision, without release (control group). Immediately after release or sham surgery, extension loss (calculated by subtracting the knee extension angle (degrees) of the operative limb from the nonoperative, contralateral limb) was measured using fluoroscopy. All animals were sacrificed following 16 weeks of postoperative free cage activity. At sacrifice, joint contracture was measured using a custom, calibrated device. The histology of the posterior joint capsule was assessed at sacrifice. All animals survived both operations without complications. Immediately after surgical release or sham surgery, the average extension loss was 129.2 ± 10.7° in the control group versus 29.6 ± 8.2° in the capsular release group (p = 0.0002). Following 16 weeks of remobilization, the average extension loss of the control and capsular release animals were 49.0 ± 12.7° and 36.5 ± 14.2°, respectively (p = 0.035). There were no histological differences between the two groups. In this animal model, a surgical capsular release decreased the extension loss (flexion contracture) immediately after surgery, as well as following sixteen weeks of remobilization. There were no histological changes detected in the posterior joint capsule. © 2013 Orthopaedic Research Society. Published by Wiley Periodicals, Inc. J Orthop Res 31:1529–1532, 2013     

兔耳瘢痕模型的应用现状

传统观念认定病理性瘢痕是人类特有的一种病理现象,不会在动物身上出现。国内外众多学者为寻求瘢痕动物模型曾作过大量的实验探索。然而,至1997年以前,还没有出现由动物自身产生的病理瘢痕实验模型。这已成为创伤研究深入发展的重大障碍。     

口服积雪苷和槲皮素对兔耳增生性瘢痕的影响

目的:探讨口服积雪苷和槲皮素对兔耳增生性瘢痕的影响。方法:新西兰大白兔9只,在兔耳腹侧制备标准创面(每耳4个,共72个),随机分为3组:空白对照组、积雪苷组及槲皮素组。4周后收集标本,从瘢痕横截面最高点制作切片,比较各组的瘢痕增生指数。结果:积雪苷组(1.55±0.03)与空白对照组(1.67±0.02)相比,瘢痕增生指数明显降低(17.9%),有统计学显著性差异(P=0.003)。槲皮素组(1.59±0.03)与空白对照组相比,两者瘢痕增生指数无统计学显著性差异(P=0.053)。结论:口服积雪苷可显著抑制兔耳瘢痕增生,口服槲皮素对兔耳增生性瘢痕的抑制作用尚待进一步实验证实。     

L-arginine reduces neointimal hyperplasia in cold-stored arterial allografts in a rabbit low-flow-through model

An "off the shelf" vascular conduit for free flap surgery that remained patent until the flap is vascularized from the recipient bed would be useful to avoid autograft donor-site morbidity and reduce operation time. To date, no successful studies of microvascular prostheses in low-flow states exist. This study investigated the effects of L-arginine on neointimal hyperplasia and the overall patency of cold-stored femoral arterial allografts implanted into a low-flow arterial defect model in rabbits. A cutaneous island flap based on the inferior epigastric artery was raised, and the femoral artery was ligated distally. Immediately proximal to the inferior epigastric artery, the femoral artery was divided and a 1-cm long cold-stored femoral arterial allograft was inserted into the defect. Half of the animals received L-arginine as a subcutaneous injection. When harvested at 4 weeks, 9 out of 11 allografts of the L-arginine-treated group and 8 out of 14 grafts of the control group were patent. Intimal thickening and graft stenosis were significantly reduced in the L-arginine group. Our findings reveal the potential of a combined approach of an arterial allograft and the use of L-arginine as a short-term vein graft substitute.     

他克莫司预防兔耳瘢痕增生的实验研究

目的：研究他克莫司对兔耳瘢痕增生的影响。方法：选10只新西兰白兔雌雄各半,按本研究组建立的改良方法制作兔耳瘢痕模型,左耳为空白对照组涂凡士林软膏,右耳为实验组涂他克莫司软膏。伤后14天、21天、28天、35天及49天采集标本,行苏木精-伊红（HE）及Masson三色法染色,统计成纤维细胞密度,实时定量PCR（Real-time PCR）检测细胞外基质成分Ⅰ型胶原（collagen Ⅰ;ColI）和纤维连接蛋白（fibronectin;Fn）,CD4＋辅助性T细胞-2（Th2）产生的重要纤维化基因IL-4及参与T细胞早期活化的重要基因巨噬细胞集落刺激因子（macrophage colony stimulating factor;M-CSF）和肿瘤坏死因子-α（tumor necrosis factorα;TNF-α）的表达。结果：HE及Masson三色法染色可见实验组较对照组胶原沉积明显减少,PCR结果可见ColI、Fn、IL-4、M-CSF和TNF-α在实验组中的表达较对照组在各时间点均减少。结论：应用他克莫司可通过下调IL-4、M-CSF和TNF-α的表达来抑制兔耳瘢痕增生。     

Acidification of formula reduces bacterial translocation and gut colonization in a neonatal rabbit model

BACKGROUND/PURPOSE: The authors hypothesized that gastric acidity is protective because it is bactericidal. They tested acidified formula for protection against gut colonization and bacterial translocation. METHODS: In vitro: Formula was acidified to pH of 2, 3, 4, 5 and innoculated with Enterobacter. Growth over time was quantitatively assessed. In vivo: 442 premature rabbit pups were sorted randomly and fed formula of pH 2, 3, 4, or 7, with ranitidine. Two models were utilized: (1) with bacterial challenge using a known acid sensitive organism, (2) without bacterial challenge to simulate natural gut colonization and to test against organisms of unknown acid sensitivity. Normal acid animals received pH 7 formula, no ranitidine. On day 3, the mesenteric lymph nodes (MLN), spleen, liver, and cecum were harvested and cultured. RESULTS: Bacterial growth was inhibited at pH 2 and 3, growth was logarithmic above pH 4 (P<.001). Total and organ-specific translocation was reduced at pH 3 and below in both models (P<.05). Translocation with formula pH 3 equaled normal acid animals. Quantitative cecal colonization was reduced in pups receiving pH 3 and below in both models (P<.05). CONCLUSION: Acidification of formula below pH 4 is bactericidal to enteric organisms. Acidified formula decreases bacterial translocation and gut colonization.     

Blocking collagen fibril formation in injured knees reduces flexion contracture in a rabbit model

Post‐traumatic joint contracture is a frequent orthopaedic complication that limits the movement of injured joints, thereby severely impairing affected patients. Non‐surgical and surgical treatments for joint contracture often fail to improve the range of motion. In this study, we tested a hypothesis that limiting the formation of collagen‐rich tissue in the capsules of injured joints would reduce the consequences of the fibrotic response and improve joint mobility. We targeted the formation of collagen fibrils, the main component of fibrotic deposits formed within the tissues of injured joints, by employing a relevant rabbit model to test the utility of a custom‐engineered antibody. The antibody was delivered directly to the cavities of injured knees in order to block the formation of collagen fibrils produced in response to injury. In comparison to the non‐treated control, mechanical tests of the antibody‐treated knees demonstrated a significant reduction of flexion contracture. Detailed microscopic and biochemical studies verified that this reduction resulted from the antibody‐mediated blocking of the assembly of collagen fibrils. These findings indicate that extracellular processes associated with excessive formation of fibrotic tissue represent a valid target for limiting post‐traumatic joint stiffness. © 2016 Orthopaedic Research Society. Published by Wiley Periodicals, Inc. J Orthop Res 35:1038–1046, 2017.

     

Inhibition of TNF-α reduces transplant arteriosclerosis in a murine aortic transplant model

Experimental and clinical data provide evidence that TNF-α contributes to acute and chronic allograft rejection. In this study, we explored the effect of TNF-α blockade using the chimeric monoclonal antibody infliximab on the development of transplant arterisoclerosis in a fully mismatched aortic allograft model. Post-transplant treatment of CBA (H2^k) recipients with 250 μg infliximab (cumulative dose 1.25 mg) reduced luminal occlusion of C57Bl/6 (H2^b) aortic grafts on day 30 from 77 ± 5% in untreated controls to 52 ± 6%. Increasing the dose of anti-TNF-α antibody had no further beneficial effect. Treatment with human control immunoglobulin had no effect on intima proliferation. Under TNF-α blockade, ICAM-1 and PDGF mRNA expression within the grafts was strongly reduced, whereas iNOS expression was enhanced. The data show that TNF-α blockade using infliximab can reduce the development of transplant arteriosclerosis in fully mismatched murine aortic grafts.     

兔耳增生性瘢痕模型建立方法的探讨

目的:探讨手术方法对兔耳瘢痕模型建立的影响。方法:建立三种兔耳皮肤创伤愈合模型,A组在切除兔耳皮肤的同时,切除皮下软骨及软骨膜;B组切除皮肤及软骨膜但保留软骨;C组只切除皮肤,保留软骨及软骨膜。对创伤愈合过程进行形态学和组织学观察,并在镜下测量瘢痕突出皮肤的高度。结果:①A、B、C三组平均创面愈合时间分别为(10.4±1.0)天、(17.8±1.6)天、(11.4±1.3)天,三组之间统计学比较有显著差异;②A、C两组伤后4～5天肉芽组织充满创面,B组在伤后第10天,肉芽组织开始在坏死软骨下向中心生长;③A、B、C三组瘢痕隆起高度为(1.20±0.56)mm、(1.68±0.73)mm、(1.15±0.50)mm,B组与A、C组统计学比较有显著差异。结论:保留软骨,切除皮肤和软骨膜能获得较大的瘢痕,可用于瘢痕治疗性实验;切除软骨形成瘢痕较小,可用于观察创面外用药物对瘢痕形成的影响。     

Probiotic fortified diet reduces bacterial colonization and translocation in a long-term neonatal rabbit model

Background

Probiotic fortified diet reduces bacterial colonization and translocation in a short-term neonatal rabbit model when continuously challenged with pathogen. The purpose of this study was to determine if live probiotic diet could remain effective at decreasing colonization/translocation of pathogens in a long-term neonatal rabbit model without ill effects of the probiotic outside the gastrointestinal (GI) tract.

Methods

Rabbit pups were born via cesarean delivery 1 day preterm and assigned to 2 diets: a newly formulated diet (controls) vs the same diet fortified with the live probiotic Lactoccocus lactis. Enterobacter cloacae was added to both preparations before each feed. Pups were gavage fed twice daily, and weights were recorded. Rabbits were sacrificed on day 7, and organs were harvested and cultured for target organism growth.

Results

The probiotic fortified diet resulted in a significant decrease in Enterobacter translocation to the liver and decreased colonization in the stomach and lungs. There was no evidence of Lactococccus translocation or colonization outside of the GI tract.

Conclusion

This probiotic fortified diet was effective at decreasing pathogenic bacteria colonization and translocation in a long-term neonatal model. The addition of L lactis to the diet resulted in appropriate growth without any colonization or translocation of the probiotic outside of the GI tract.