1、Designation: F3207 17Standard Guide forin vivo Evaluation of Rabbit Lumbar Intertransverse ProcessSpinal Fusion Model1This standard is issued under the fixed designation F3207; the number immediately following the designation indicates the year oforiginal adoption or, in the case of revision, the ye
2、ar of last revision. A number in parentheses indicates the year of last reapproval. Asuperscript epsilon () indicates an editorial change since the last revision or reapproval.1. Scope1.1 Historically, the single-level rabbit posterolateral, orintertransverse, lumbar spine fusion model was developed
3、 andreported on by Dr. Scott Boden, et. al. (Emory Spine Center forOrthopedics) and the model has been proposed as a non-clinical model which may be used to replicate clinically-relevant fusion rates for iliac crest autograft in the posterolat-eral spine (1, 2).2This model is used routinely in submi
4、ssionsto regulatory bodies for the purpose of evaluating the potentialefficacy of bone void filler materials as compared to othermaterials or iliac crest autograft to effect spinal posterolateralfusion. The use of this standards recommendations as part ofa regulatory submission does not provide any
5、guarantee ofregulatory clearance and should be considered as a part of thedata provided for regulatory submission.1.2 This guide covers general guidelines to evaluate theeffectiveness of products intended to cause and/or promotebone formation in the lumbar intertransverse process spinalfusion model
6、in vivo. This guide is applicable to products thatmay be composed of one or more of the following components:natural biomaterials (such as demineralized bone), and syn-thetic biomaterials (such as calcium sulfate, glycerol, andreverse phase polymeric compounds) that act as additives,fillers, and/or
7、excipients (radioprotective agents, preservatives,and/or handling agents). It should not be assumed that productsevaluated favorably using this guidance will form bone whenused in a clinical setting. The primary purpose of this guide isto facilitate the equitable comparison of bone void fillers and/
8、orautograft extender products in vivo. The purpose of this guideis not to exclude other established methods.1.3 The values stated in SI units are to be regarded asstandard. No other units of measurement are included in thisstandard.1.4 This standard does not purport to address all of thesafety conce
9、rns, if any, associated with the use of bone voidfillers. It is the responsibility of the user of this standard toestablish appropriate safety and health practices involved inthe development of said products in accordance with appli-cable regulatory guidance documents and in implementing thisguide t
10、o evaluate the bone-forming/promoting capabilities ofthe product.1.5 This standard does not purport to address the require-ments under 21 CFR Part 58 concerning Good LaboratoryPractices or international standard counterpart OECD Prin-ciples of Good Laboratory Practice (GLP). It is the responsi-bilit
11、y of the sponsor of the study to understand the require-ments for conduct of animal studies whereby the data may beused to support premarket applications, including require-ments for personnel, protocol content, record retention andanimal husbandry.1.6 This international standard was developed in ac
12、cor-dance with internationally recognized principles on standard-ization established in the Decision on Principles for theDevelopment of International Standards, Guides and Recom-mendations issued by the World Trade Organization TechnicalBarriers to Trade (TBT) Committee.2. Referenced Documents2.1 A
13、STM Standards:3E122 Practice for Calculating Sample Size to Estimate, WithSpecified Precision, the Average for a Characteristic of aLot or ProcessE1402 Guide for Sampling DesignE1488 Guide for Statistical Procedures to Use in Developingand Applying Test MethodsF2529 Guide forin vivo Evaluation of Os
14、teoinductive Poten-tial for Materials Containing Demineralized Bone (DBM)F2884 Guide for Pre-clinical in vivo Evaluation of SpinalFusion2.2 Federal Documents:421 CFR 58 Good Laboratory Practice for Nonclinical Labo-ratory Studies1This guide is under the jurisdiction of ASTM Committee F04 on Medical
15、andSurgical Materials and Devices and is the direct responsibility of SubcommitteeF04.44 on Assessment for TEMPs.Current edition approved June 1, 2017. Published July 2017. DOI: 10.1520/F3207-172The boldface numbers in parentheses refer to the list of references at the end ofthis standard.3For refer
16、enced ASTM standards, visit the ASTM website, www.astm.org, orcontact ASTM Customer Service at serviceastm.org. For Annual Book of ASTMStandards volume information, refer to the standards Document Summary page onthe ASTM website.4Available from U.S. Food and Drug Administration (FDA), 10903 NewHamps
17、hire Ave., Silver Spring, MD 20993, http:/www.fda.gov.Copyright ASTM International, 100 Barr Harbor Drive, PO Box C700, West Conshohocken, PA 19428-2959. United StatesThis international standard was developed in accordance with internationally recognized principles on standardization established in
18、the Decision on Principles for theDevelopment of International Standards, Guides and Recommendations issued by the World Trade Organization Technical Barriers to Trade (TBT) Committee.12.3 AAMI/ISO Documents:5ISO 10993-6 Third edition 2016-12-01 Biological evalua-tion of medical devices Part 6: Test
19、s for local effectsafter implantation3. Terminology3.1 Definitions:3.1.1 biomechanical fusion, nthe increased strengthand/or stiffness and reduced ROM of a spinal unit as comparedto that measured before surgical intervention.3.1.2 biomechanical properties, nas used in thisdocument, evaluation of the
20、 operative functional spinal unitmultidirectional range of motion (ROM: Lateral bending,Flexion Extension and Axial Rotation) properties undernon-destructive conditions, tensile stiffness and ultimate load.3.1.3 fusion, na multifactorial outcome which can becharacterized in terms of the radiographic
21、, biomechanical andhistological results of the intended spinal arthrodesis proce-dure.3.1.4 histological evidence of fusion, nbased on lightmicroscopy of newly formed and remodeled bone spanning theintertransverse region, with contiguous osseous connectivityobserved between the adjacent transverse p
22、rocesses. Assess-ment rationale must be justified.3.1.5 manual palpation, na method for evaluating spinalfusion status by estimating the stiffness of the operative motionsegment and adjacent superior motion segment by the appli-cation of multidirectional loads in lateral bending and flexion-extensio
23、n using the hands.3.1.6 micro-computed tomographic (micro-CT) fusion,ntomographic fusion is based on interpretation of themicro-CT images, with fusion success based on the three-dimensional appearance of contiguous bone from transverseprocess to transverse process (i.e. bridging bone).3.1.7 non-unio
24、n, na multifactorial outcome which can becharacterized in terms of the radiographic, biomechanical andhistological results indicating a lack of trabecular or corticalbone spanning the intertransverse region, without contiguousosseous connectivity observed between the adjacent transverseprocesses.3.1
25、.8 radiographic fusion, nstatus of radiographic fusionis based on interpretation of the posteroanterior (P/A) plainfilm x-ray images, with fusion success based on the appearanceof contiguous bone from transverse process to transverseprocess (i.e. bridging bone).4. Significance and Use4.1 This guide
26、covers animal implantation methods andanalysis of bone void fillers to determine whether a material orsubstance leads to lumbar intertransverse process spinal fusion,as defined by its ability to cause bone to form in vivo.5. Animal Models5.1 General NoteAppropriate positive or comparativecontrols ma
27、y be used. For example, comparative controls couldbe similar devices, and positive controls could be autograftfrom the animal.5.2 Skeletally mature New Zealand white rabbits(typically 7 months and 3.5-4.5kg). Proximal tibial and distalfemoral physes should be closed and verified via plain radio-grap
28、hs. Radiographic and histologic closure of the distalfemoral growth plates occur at an average age of 21 and 22weeks, respectively. The proximal tibial physes close radio-graphically and histologically at an average age of 26 and 28weeks, respectively. A lateral radiograph is a more reliablemethod f
29、or assessing physeal closure in the rabbit, and radio-graphic confirmation of tibial physeal closure should beobtained prior to using rabbits that are younger than approxi-mately 7 months of age. Radiographic confirmation of physealclosure is probably not necessary in rabbits 8 months or olderbut sh
30、ould be provided for the sake of completeness. Someminor variation in age of tibial growth plate closure may beexpected with different strains of New Zealand White rabbits.Weight is not a reliable indicator of skeletal maturity in theNew Zealand White rabbit. All rabbits used should be of thesame se
31、x. (3, 4)5.3 Implant Mass/VolumeIn general, implant mass (1.6-2.2 grams; useful only for autograft)/volume (2.5-3.0 cc) perside is used. It is recommended that the experimental groupcontain the same total implant volume as any comparativegroups so the results are comparable and the potential effects
32、 ofthe implant on intertransverse process spinal fusion can bedetermined.5.4 Sample Size:5.4.1 Sample sizes should be justified in the study protocoland, if possible, should provide statistical power appropriate tothe endpoint using appropriate statistical methods to justify asrequired. Interim time
33、 points may be used as appropriate andjustifications should be provided. Should statistical numbersnot be practical or possible, empirical testing in the literaturehas shown an n=6-8 to be a target sample size minimum.5.4.2 Sample size should be determined with reference tothe primary outcome of the
34、 study, which is typically the fusionrate at 8 or 12 weeks. Additionally, it may be necessary toconsider the sampling requirements of other analyses in thestudy; in particular, quantitative endpoints such as morphom-etry.NOTE 1The sample size recommendations refer to the number ofsamples expected to
35、 be available for analysis.Attrition, or loss of animalsdue to surgical complications, is common in the rabbit spinal fusion model(especially with autograft harvesting). It may be necessary to plan foradditional animals to replace those lost to attrition. Make sure you reportall animals treated, any
36、 unexpected or early deaths, etc.5.5 End PointsEach implant group should have an imme-diate post-operative assessment and end points should bejustified by the resorption profile of the materials; there shouldbe at least 2 time points less than the maximum assessmenttime (an early and mid-phase) in o
37、rder to assess any irregu-larities (unexpected or excessive inflammation, etc.) at the5Available from Association for the Advancement of Medical Instrumentation(AAMI), 4301 N. Fairfax Dr., Suite 301, Arlington, VA 22203-1633, http:/www.aami.org.F3207 172implant and peri-implant site (recommended tim
38、e periodsrepresentative in the literature are 4, 8, and 12 weeks) or longerperiods may be warranted and should be justified.6. Recommended Surgical Protocol Methodologies6.1 Rabbit Lumbar Intertransverse Process Spinal FusionRecommended Surgical Technique:6.1.1 Aseptic technique should be employed d
39、uring thesurgical implantation procedures.6.1.2 Animals should be singly housed in standard cagesand fed with rabbit food and water.NOTE 2Handling of the animals during the first 14 days post-opshould be avoided unless medically required.6.1.3 Pre-operative analgesics: 0.05 mg/ kg buprenorphineadmin
40、istered subcutaneously and the application of a fentanylpatch (25 g/hr) to the inner ear pinna, or other analgesicapproved by the IACUC. A 25 g/hr fentanyl patch is aneffective analgesic with duration of up to 72 hours, but mayrequire up to 12 hours after application until blood levels aresufficient
41、 to provide pain relief. Patches may be placed theevening prior to surgery or animals dosed with an analgesicsuch as butorphanol prior to surgery and several hours aftersurgery to ensure analgesisc coverage while fentanyl bloodlevels rise.6.1.4 Anesthetics: Induction and maintenance: 34 mg/ kgketami
42、ne and 5 mg/ kg xylazine administered intramuscularly.Isoflurane should be administered via laryngeal mask within arange of 2%-3%, but increasing and decreasing the percentageadministered should be based on the individual animal re-sponse. Ophthalmic ointment should be applied to the eyesfollowing p
43、re-anesthesia and prior to surgery.6.1.5 Identify each animal with a unique identifier (ear tag,tattoo, etc.). Record the individual animal identification num-bers along with the body weights.6.1.6 Sedate the animal with an IACUC approved medica-tion and maintain general anesthesia with Isoflurane o
44、r otheranesthetic approved by the IACUC. The depth of anesthesiashould be sufficient to prevent muscular movement. This canbe checked by pinching the toe (between the digits) of theanimals hind limbs. If there is a reflex reaction, the animal isnot sufficiently anesthetized to continue with the impl
45、antation.Atechnician shall monitor the animals vitals/parameters whileunder anesthesia and record every 15 minutes.6.1.7 Place the anesthetized animal in a sternal or ventralrecimbant position on a clean flat surface in a procedure roomand shave the dorsum of the animal from the mid thoracicregion w
46、ell below the iliac crests with clippers. Scrub theclipped area with surgical scrub (chlorhexidine scrub or povi-done scrub). Start from the center and work, in a circularfashion, to the edge of the surgical area. Wipe off the surgicalscrub with 70 % isopropyl alcohol (repeat entire scrub proce-dure
47、 at least 3 times). The surgeon will complete finalpreparation for aseptic surgery.6.1.8 Transfer the anesthetized animal to the surgical suite.6.1.9 Lumbar Posterolateral Intertransverse spinal fusion isdetailed as follows:6.1.9.1 Final sterile prep of the surgical site is completed inthe operating
48、 room with 2% chlorhexidine or povidone solu-tion prior to first incision. Start from the center and work to theedge of the surgical area. Wipe off the solution with a clean,sterile gauze pad. The spinal level to be fused, most commonlyL4L5 or L5L6, is then identified by palpation. A line drawnfrom
49、the most cranial aspect of one iliac crest to the other, theintercrestal line, will generally pass between the L6 and L7spinous processes (Fig. 1). A second method to verify thecorrect operative level is based on the anatomy of the lum-bosacral spinous processes. Specifically, there is often a muchwider interspinous distance at L6L7 than there is at L5L6,L7S1 or between the sacral processes (Fig. 1), although this isnot always the case.6.1.9.2 Using both techniques of localization, the L4L5 orL5L6 level can be correctly identified in the vast majority
