Abstract:
Methods for repairing a cartilage defect in a subject, such methods comprising placing a tissue specimen into a container, centrifuging the container to separate the specimen into at least three fractions, drawing a selected fraction from the container, processing the fraction into a therapeutic composition, and treating the cartilage defect with the therapeutic composition.

Description:
INTRODUCTION 
       [0001]    The present technology relates to methods, compositions, and devices for repairing cartilage defects. 
         [0002]    Cartilage defects can appear as a hole or a tear in a cartilage surface and can result from wear, trauma or disease. Since cartilage has minimal ability to repair itself, even a small cartilage defect, if left untreated, can hinder a person&#39;s ability to move free from pain and can cause deterioration of a joint surface. Traditional treatments for cartilage defects include trimming the defect from the surface of the cartilage using orthoscopic surgery, or repairing the defect with sutures. More recently, treatments for cartilage defects have included the harvesting of cartilage cells, which are then cultured and implanted back into the cartilage defect to regenerate cartilage. 
       SUMMARY 
       [0003]    The present technology provides methods for repairing a cartilage defect in a human or animal subject. Such methods include a method for treating a cartilage defect comprising: obtaining blood compatible with the subject; fractionating the blood to produce platelet-poor plasma; concentrating the platelet-poor plasma to produce a platelet-poor plasma concentrate; and administering the concentrate to the site of the cartilage defect. The blood may be obtained from the subject and fractionated by centrifuging the blood to form platelet-poor plasma. The centrifuging may be performed using a container including a buoy that is able to separate the blood into two or more fractions having different densities. 
         [0004]    Some methods further comprise administering to the cartilage defect an adjunct therapeutic material. The adjunct therapeutic material may be selected from the group consisting of bioactive agents, scaffold materials, isolated tissue materials, and combinations thereof. Bone marrow aspirate is used as a therapeutic material in some methods. 
         [0005]    Further areas of applicability will become apparent from the description provided herein. It should be understood that the description and specific examples are intended for purposes of illustration only and are not intended to limit the scope of the present technology. 
     
    
     
       DRAWINGS 
         [0006]    The present technology will become more fully understood from the detailed description and the accompanying drawings, wherein: 
           [0007]      FIG. 1  illustrates a representative site of a cartilage defect in a subject in need of treatment according to some embodiments of the present technology; 
           [0008]      FIG. 2  is a diagrammatic illustration of a representative method for treating a cartilage defect according to one embodiment of the present technology; 
           [0009]      FIG. 3  is a cross-sectional view of the representative device used for isolating a blood component according to one embodiment of the present technology; 
           [0010]      FIGS. 4A and 4B  are cross-sectional views of a representative device used for forming a therapeutic composition according to one embodiment of the present technology; 
           [0011]      FIG. 5  illustrates a representative manner of administrating a cartilage defect treatment to the subject according to one embodiment of the present technology; and 
           [0012]      FIG. 6  is a perspective view of a kit useful for treating a cartilage defect according to one embodiment of the present technology. 
       
    
    
     DETAILED DESCRIPTION 
       [0013]    The following description of technology is merely exemplary in nature of the subject matter, manufacture, and use of one or more inventions, and is not intended to limit the scope, application, or uses of any specific invention claimed in this application or in such other applications as may be filed claiming priority to this application, or patents issuing therefrom. 
         [0014]      FIG. 1  is an example of a cartilage defect  130  in a human subject  100 . The cartilage defect  130  is in a knee which comprises a femur  102 , a tibia  103 , a fibula  105 , a patella  106 , and cartilage  120 . It should be understood, however, that the cartilage defect may be in any joint of a human subject  100  or animal subject, including shoulders, elbows, wrists, ankles, hips, and the spinal column, in which cartilage tissue is inadequate for physiological or cosmetic purposes. In this regard, cartilage defects include congenital cartilage defects, cartilage defects that result from or are symptomatic of disease, disorder, or trauma, and cartilage defects that are consequent to surgical or other medical procedures. For example, cartilage defects may be defects resulting from osteoporosis, spinal fixation procedures, hip and other joint replacement procedures, and chronic wounds. 
         [0015]    One embodiment for treatment of a cartilage defect  130  is shown diagrammatically in  FIG. 2 . In summary, platelet-poor plasma is obtained in step  14 . The platelet-poor plasma is then processed in step  16  to form a therapeutic composition comprising concentrated platelet-poor plasma. An adjunct therapeutic material may also be combined with the concentrated platelet-poor plasma in step  18 . The therapeutic composition from step  16  is then administered to a cartilage defect  130  in step  20 , as further shown in  FIG. 5 . Each of the aforementioned steps will be more fully discussed below. 
         [0016]    As discussed above, platelet-poor plasma is obtained at step  14 . The platelet-poor plasma is preferably isolated from blood obtained from the subject  100  exhibiting the cartilage defect  130  to be treated. The blood may also be bone marrow derived. Platelet-poor plasma can be isolated in step  14  by a variety of methods, including by density fractionation of blood, cryopreciptation, and filtration. Density fractionation includes single stage centrifugation, centrifugation in multiple stages, and continuous flow centrifugation. 
         [0017]      FIG. 3  illustrates one example of a separation device that can be used for forming the platelet-poor plasma in step  14  by density fractionation. In this regard, the device  22  includes a container  24 , such as a tube, that is placed in a centrifuge after being filled with blood. The container  24  includes a buoy system having an isolator  26  and a buoy  28 . The buoy  28  has a selected density which is tuned to reach a selected equilibrium position upon centrifugation; this position lies between a more dense blood fraction and a less dense blood fraction. During centrifugation, the buoy  28  separates the blood within the container  24  into at least two fractions, without substantially commingling the fractions, by sedimenting to a position between the two fractions. In this regard, the isolator  26  and the buoy  28  define a layer comprising platelet-rich plasma  30 , while less dense platelet-poor plasma  32  generally fractionates above the isolator  26 , and more dense red blood cells  34  generally fractionate below the buoy  28 . Following centrifugation, a syringe or tube may then be interconnected with a portion of the buoy system to extract one or more selected fractions for use as the blood component. Devices including those disclosed in  FIG. 3  and associated methods are described in U.S. Patent Application Publication 2004/0251217, Leach et al., published Dec. 12, 2004; and U.S. Patent Application Publication 2005/0109716, Leach et al., published May 26, 2005; both of which are incorporated by reference herein. One such device that is commercially available is the GPS® Platelet Concentrate System, from Biomet Biologics, Inc. (Warsaw, Ind.). 
         [0018]    The platelet-poor plasma obtained in step  14  is processed into a therapeutic composition comprising concentrated platelet-poor plasma in step  16 . One example of a concentration device  40  that may be used for forming concentrated platelet-poor plasma in step  16  is shown in  FIGS. 4A and 4B . In this regard, the concentration device  40  has an upper chamber  41  and a lower chamber  42 . The upper chamber  41  has an end wall  43  through which the agitator stem  44  of a gel bead agitator  45  extends. The concentration device  40  also has a plasma inlet port  46  that extends through the end wall  43  and into the upper chamber  41 . The concentration device  40  also includes an outlet port  47  that communicates with a plasma concentrate conduit  48 . The floor of upper chamber  41  includes a filter  49 , the upper surface of which supports desiccating gel beads  50 . 
         [0019]    During use, platelet-poor plasma  52 , with or without optional materials such as an adjunct therapeutic material discussed below, is introduced into the upper chamber  41  through the plasma inlet port  46 . The platelet-poor plasma  52  flows to the bottom of the upper chamber  41  where it contacts the polyacrylate beads  50  as shown in  FIG. 4A . As the polyacrylate beads  50  remove water from the platelet-poor plasma  52 , the plasma  52  thickens. During this concentration stage, the platelet-poor plasma  52  and its components can be concentrated to a concentration of from about 1.5 to 3 times or higher than its original concentration to create the therapeutic composition  53 . 
         [0020]    Referring to  FIG. 4B , the concentration device  40  is then placed in the cup receptors of a conventional laboratory centrifuge (not shown) and spun at a speed that will create a centrifugal force that will remove the therapeutic composition  53  from the polyacrylate gel beads  50 , and cause the therapeutic composition  53  to flow through the filter  49 . The filter  49  can be constructed to allow flow of liquid there-through at centrifugal forces above 10 g. After centrifugation is completed, the concentration device  40  is removed from the centrifuge. The platelet-poor plasma therapeutic composition  53  is then drawn from the lower chamber  42  through the conduit  48  to the outlet port  47 . In some embodiments, the therapeutic composition  53  forms a gel. 
         [0021]    Exemplary plasma concentration devices are disclosed in U.S. Patent Application Publication 2006/0175268, Dorian et al., published Aug. 10, 2006; and U.S. Patent Application Publication 2006/0243676, Swift et al., published Nov. 2, 2006; both of which are incorporated by reference herein. Such a device is commercially available as Plasmax™ Plus Plasma Concentrator, from Biomet Biologics, Inc. (Warsaw, Ind.). 
         [0022]    The therapeutic composition obtained in step  16  may include optional materials that are combined with concentrated platelet-poor plasma in step  18 . Optional materials include, for example, adjunct therapeutic materials such as platelet activators or other bioactive agents, scaffolds, buffers, isolated tissue materials and combinations thereof. Such adjunct therapeutic materials may be added to platelet-poor plasma prior to concentration of the platelet-poor plasma in step  16 , or may be added to the therapeutic composition after concentration of the platelet-poor plasma in step  16 . 
         [0023]    Isolated tissue materials useful as optional materials in step  18  comprise tissue material that has been extracted from a human or other animal subject and which, in some embodiments, has been subjected to processing prior to mixing with concentrated platelet-poor plasma. Examples of isolated tissue material include platelet-rich plasma or other blood component, bone marrow aspirate, concentrated bone marrow aspirate, and processed lipoaspirate cells. The isolated tissue material may contain hematopoietic stem cells, stromal stem cells, mesenchymal stem cells, endothelial progenitor cells, red blood cells, white blood cells, fibroblasts, reticulocytes, adipose cells, thrombocytes, and endothelial cells. The isolated tissue material may be autologous tissue, i.e., tissue from the subject  100  having the cartilage defect  130  to be treated. 
         [0024]    The isolated tissue material of step  18  may comprise bone marrow aspirate or concentrated bone marrow aspirate. Bone marrow aspirate can be obtained in any appropriate manner, such as from the intramedullary area of a bone by use of a syringe and needle. The bone marrow aspirate may be used as-is in step  18 , or may be further processed to create bone marrow concentrate or other isolated tissue composition. In some embodiments, a separation device, such as shown in  FIG. 3 , may be used to obtain a concentrated bone marrow aspirate comprising nucleated cells, such as red and white blood cells, bone marrow stromal cells, and mesenchymal stem cells. For example, a mixture of whole blood and bone marrow aspirate may be added to the separation device  22  shown in  FIG. 3 , and a buffy coat fraction (platelet-rich plasma  30 ) obtained that contains at least a 4 times greater concentration of nucleated cells from bone marrow. Methods of obtaining an isolated tissue composition from bone marrow aspirate are disclosed in U.S. Patent Application Publication No. 2006/0278588 to Woodell-May published Dec. 14, 2006, which is incorporated by reference herein. 
         [0025]    Other devices that may be used to obtain the isolated tissue composition at step  18  are described, for example, in U.S. Pat. No. 6,398,972, Blasetti et al., issued Jun. 4, 2002; U.S. Pat. No. 6,649,072, Brandt et al., issued Nov. 18, 2003; U.S. Pat. No. 6,790,371, Dolecek, issued Sep. 14, 2004; U.S. Pat. No. 7,011,852, Sukavaneshvar et al., issued Mar. 14, 2006; U.S. Patent Application Publication 2005/0196874, Dorian et al., published Sep. 8, 2005; and U.S. Patent Application Publication 2006/0175242, Dorian et al., published Aug. 10, 2006. In addition to the GPS® Platelet Concentrate System, a variety of other commercially available devices may be used to obtain the isolated tissue composition at step  18 , including the Megellan™ Autologous Platelet Separator System, commercially available from Medtronic, Inc. (Minneapolis, Minn.); SmartPReP™, commercially available from Harvest Technologies Corporation (Plymouth, Mass.); DePuy (Warsaw, Ind.); the AutoloGel™ Process, commercially available from Cytomedix (Rockville, Md.), and the Genesis CS component concentrating system, available from EmCyte Corporation (Fort Myers, Fla.). 
         [0026]    The isolated tissue composition of step  18  may comprise stem cells, such as bone marrow-derived stem cells and adipose-derived stromal cells. Adipose-derived stromal cells may be obtained from processing of lipid tissue by standard liposuction and lipaspiration methods known in the art. Adipose tissue may also be treated with digestive enzymes and with chelating agents that weaken the connections between neighboring cells, making it possible to disperse the tissue into a suspension of individual cells without appreciable cell breakage. Following disaggregation, the adipose stromal cells may be isolated from the suspension of cells and disaggregated tissue. A device such as the GPS® Platelet Concentrate System, may be used to isolate adipose stromal cells. 
         [0027]    Platelet activators optionally included in step  18  may serve to activate one or more growth factors within platelets that optionally may be in the therapeutic composition. Activation of the platelets by the platelet activators can be performed just prior to administration of the therapeutic composition, concomitant with administration of the therapeutic composition, or following administration of the therapeutic composition to the cartilage defect in step  20 . Platelet activators among those useful herein include thrombin, calcium chloride (CaCl 2 ), coagulation factors, and mixtures thereof. Coagulation factors include, but are not limited to, one or more of the following: V, VII, VIIa, IX, IXaβ, X, Xa, XI, XIa, XII, α-XIIa, β-XIIa, and XIII. 
         [0028]    A scaffold may be added in step  18  to contain, support, or retain the therapeutic composition at the cartilage defect site, or to facilitate migration of endogenous cells into the administration site. Scaffolds may be formed from porous or semi-porous, natural, synthetic or semisynthetic materials. Scaffold materials include those selected from the group consisting of bone (including cortical and cancellous bone), demineralized bone, ceramics, polymers, and combinations thereof. Bone, demineralized bone and ceramics may be particularly useful in methods where the therapeutic composition is applied to subchondral bone, as in a microfracture procedure. Suitable polymers may include collagen, including lyophilized or skin-derived collagen as disclosed in U.S. patent application Ser. No. 11/259,216 which is incorporated by reference herein. Polymers may also include gelatin, hyaluronic acid, chitosan, polyglycolic acid, polylactic acid, polypropylenefumarate, polyethylene glycol, and copolymers or combinations thereof. Ceramics include any of a variety of ceramic materials known in the art for use for implanting in bone, such as calcium phosphate (including tricalcium phosphate, tetracalcium phosphate, hydroxyapatite, and mixtures thereof). 
         [0029]    Referring again to  FIG. 2 , the therapeutic composition  53  created in step  16  is administered to the cartilage defect  130  in step  20  according to any medically appropriate procedure. For example, as noted above, a microfracture procedure may be performed at the site of the cartilage defect  130  prior to administering the therapeutic composition in step  20 . In such a method, the subchondral bone adjacent to the cartilage defect is breached, and the concentrate is administered to the site of the breach. 
         [0030]    As illustrated in  FIG. 5 , a dual syringe device  200  may be employed in a medically appropriate procedure. The dual syringe device  200  includes a first barrel  201  and a second barrel  202 , both of which are connected to a mixing chamber  212 . A first plunger  205  is inserted into the first barrel  201  and a second plunger  206  is inserted into the second barrel  202 . The first plunger  205  and the second plunger  206  are connected by a member  208 . The mixing chamber  212  connects to a cannula  215 . In some embodiments, the dual syringe device  200  contains concentrated platelet-poor plasma  53  in the first barrel  201 , and an adjunct therapeutic material  260 , such as a platelet activator, in the second barrel  202 . During step  20  of administering the therapeutic composition, member  208  is pushed toward the mixing chamber  212  such that the contents of both the first barrel  201  and the second barrel  202  are pushed into the mixing chamber  212 . The therapeutic composition  250  in the mixing chamber  212  is pushed through the cannula  215  onto the cartilage defect  130 . In some embodiments, depending on the adjunct therapeutic material  260 , the therapeutic composition  250  can form a clot. 
         [0031]    In some embodiments, the dual syringe device  200  is used to pierce soft tissue of the subject  100  to repair the cartilage defect  130 . An incision may be made in the subject  100  to allow entry of the cannula  215  so that the dual syringe device  200  may enter an area of the cartilage defect  130 . 
         [0032]    The present technology also provides a cartilage repair system comprising a consumable component of a separation device, such as the separation device  22  illustrated in  FIG. 3 , and a concentration device  40  illustrated in  FIGS. 4A and 4B . The cartilage repair system can also include a surgical process component operable to facilitate treatment of the cartilage defect  130  in the subject  100 . 
         [0033]    The present technology also provides kits to facilitate the methods described herein. As illustrated in  FIG. 6 , a kit  300  comprises one or more components, materials or devices used in such methods. A kit  300  can be placed in a tray  302  which is to provide a clean and sterile environment for use of the kit&#39;s contents during a method of the present technology. 
         [0034]    The kit  300  may include a separation device  22  and a concentration device  40  such as illustrated in  FIG. 3  and  FIG. 4 , respectively. The kit  300  also includes, for example, a first syringe  329  (e.g., with a 60 ml capacity) and a needle  327  to draw blood from the subject  100 . The kit  300  may also include an anticoagulant solution  330 , which may be drawn into first syringe  329  prior to drawing blood from the subject  100 . The first syringe  329  can also be used to inject blood into the separation device  22 . The kit may also contain a second syringe  328  (e.g., with a 30 ml capacity) for use in extracting platelet-poor plasma from the separation device  22  after centrifuging the separation device  22  with the blood. The platelet-poor plasma in the second syringe  328  may be injected into the concentration device  40 . The kit may contain a third syringe  324  (e.g., with a 10 ml capacity) for use in withdrawing platelet-rich plasma from the separation device  22  if platelet-rich plasma is desired for use as an adjunct therapeutic material. The kit further contains a fourth syringe  325  (e.g., having a capacity of 10 ml) for extracting the therapeutic composition containing platelet-poor plasma from concentration device  40 . 
         [0035]    The kit  300  can further include an infusion cannula  314  that may be used for administering the therapeutic composition, as well as other materials and devices to facilitate the methods of the present technology. For example, a tourniquet  320 , gauze  316 , tape  318 , antiseptic wipes  322 , or other medical supplies may be provided to assist the practitioner. In some embodiments, the kit  300  can include a dual syringe device  200  such as illustrated in  FIG. 5 . The kit  300  can also include an adjunct therapeutic material such as a platelet activator, or an anticoagulant  330  as discussed above. 
         [0036]    The systems and kits of the present technology may also include means of communicating information and/or instructions. The communication means may include language as required by an organization or government agency such as the United States Food &amp; Drug Administration. The communication means can include labels; package inserts; brochures; advertisements; computer readable digital optical media, for example, diskettes or CD&#39;s; audio or video presentations, for example, audio tapes, CD&#39;s, or DVD&#39;s, and/or one or more pages on a website. 
         [0037]    The embodiments and the examples described herein are exemplary and not intended to be limiting in describing the full scope of the devices, compositions and methods of the present technology. Equivalent changes, modifications and variations can be made within the scope of the present technology, with substantially similar results.