Name: preparation, procedures and evaluation of platelet-rich plasma injection in the treatment of knee osteoarthritis
Uploaded: 2019-01-04T13:00:00
Description: Watch this Scientific Journal Video about Preparation, Procedures and Evaluation of Platelet-Rich Plasma Injection in the Treatment of Knee Osteoarthritis at JoVE.com

This study was supported by the National Natural Science Foundation of China (81371991) and the Major Program of Science and Technology of Guangdong (2015B020225007).

The methods described were approved by the Ethics Committee of Guangdong General Hospital.
1. Obtain PRP by Manual Centrifugation
<ol>
	<li>Prepare the patient in a supine position in a sterile laminar flow operating room with a comfortable room temperature and humidity: room temperature is 22 &#176;C and room humidity is 60%.</li>
	<li>Use a 1-mL syringe to draw 0.2 mL of heparin sodium (2 mL = 12,500 U), and then moisten a 50-mL syringe. 
	NOTE: 3 mL of sodium citrate is also typical ...

<ol>
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P. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Radiofrequency+procedures+to+relieve+chronic+knee+pain:+an+evidence-based+narrative+review.">Radiofrequency procedures to relieve chronic knee pain: an evidence-based narrative review.</a> Reg Anesth Pain Med. 43 (1), 72 (2018).</li><li>S&#225;nchez, M., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=A+randomized+clinical+trial+evaluating+plasma+rich+in+growth+factors+(PRGF-endoret)+versus+hyaluronic+acid+in+the+short-term+treatment+of+symptomatic+knee+osteoarthritis.">A randomized clinical trial evaluating plasma rich in growth factors (PRGF-endoret) versus hyaluronic acid in the short-term treatment of symptomatic knee osteoarthritis.</a> Arthroscopy the Journal of Arthroscopic &amp; Related Surgery. 28 (8), 1070-1078 (2012).</li><li>LaPrade, R. F., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Biologic+treatments+for+sports+injuries+II+think+tank-current+concepts,+future+research,+and+barriers+to+advancement,+part+1:+biologics+overview,+ligament+injury,+tendinopathy.">Biologic treatments for sports injuries II think tank-current concepts, future research, and barriers to advancement, part 1: biologics overview, ligament injury, tendinopathy.</a> Am J Sports Med. 44 (12), 3270-3283 (2016).</li><li>Ornetti, P., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Does+platelet-rich+plasma+have+a+role+in+the+treatment+of+osteoarthritis?.">Does platelet-rich plasma have a role in the treatment of osteoarthritis?.</a> Joint Bone Spine. 83 (1), 31-36 (2016).</li><li>Knop, E., Paula, L. E. D., Fuller, R. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Platelet-rich+plasma+for+osteoarthritis+treatment.">Platelet-rich plasma for osteoarthritis treatment.</a> Revista Brasileira de Reumatologia (English Edition). 56 (2), 152-164 (2016).</li><li>Weibrich, G., Kleis, W. K., Hafner, G., Hitzler, W. E. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Growth+factor+levels+in+platelet-rich+plasma+and+correlations+with+donor+age,+sex,+and+platelet+count.">Growth factor levels in platelet-rich plasma and correlations with donor age, sex, and platelet count.</a> J Craniomaxillofac Surg. 30 (2), 97-102 (2002).</li><li>Landesberg, R., Roy, M., Glickman, R. S. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Quantification+of+growth+factor+levels+using+a+simplified+method+of+platelet-rich+plasma+gel+preparation.">Quantification of growth factor levels using a simplified method of platelet-rich plasma gel preparation.</a> Journal of Oral &amp; Maxillofacial Surgery. 58 (3), 297-300 (2000).</li><li>Wasterlain, A. S., Braun, H. J., Dragoo, J. L. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Contents+and+formulations+of+platelet-rich+plasma.">Contents and formulations of platelet-rich plasma.</a> Operative Techniques in Orthopaedics. 22 (1), 33-42 (2012).</li><li>Gobbi, G., Vitale, M. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Platelet+rich+plasma+for+biological+therapy:+applications+and+limits.">Platelet rich plasma for biological therapy: applications and limits.</a> Operative Techniques in Orthopaedics. 22 (1), 10-15 (2012).</li><li>Chahla, J., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=A+call+for+standardization+in+platelet-rich+plasma+preparation+protocols+and+composition+reporting:+a+systematic+review+of+the+clinical+orthopaedic+literature.">A call for standardization in platelet-rich plasma preparation protocols and composition reporting: a systematic review of the clinical orthopaedic literature.</a> Journal of Bone &amp; Joint Surgery-american. 99 (20), 1769-1779 (2017).</li><li>Delong, J. 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The concentration of blood platelets in normal human blood is between 150,000/&#181;L and 350,000/&#181;L, and it is widely believed that blood platelet concentration of PRP should reach 1,000,000/&#181;L, which is 3 to 5 times normal concentrations9. According to the PAW hierarchy system, it is believed that PRP has no obvious effect when the blood platelet concentration is less than three times the normal concentration, while PRP has an inhibiting effect when its blood platelet concentration is ...

Knee osteoarthritis (KOA) is one of the most frequently seen diseases in the orthopedic department; 30%-50% of people over the age of 65 years experience this disease1. At present, the conservative management of KOA mainly includes oral administration of non-steroidal anti-inflammatory drugs and cartilage nutrient drugs, intra-articular injection of sodium hyaluronate, and physiotherapy. However, these methods cannot stop the process of knee joint degeneration2. Articular cartilage defects can cause articular surface wear, joint instability, and metabolic changes, which are part of the pathogenesis of KOA3. However, because of the absence of blood vessels, nerves and lymphoid tissue in articular cartilage, recovery after damage is difficult. An effective method of repairing cartilage is especially important for the treatment of KOA. The treatment of osteoclasia is also a key focus in KOA treatment.
Platelet-rich plasma (PRP) is an autologous bioactive substance, and the application of PRP to bone and joint problems is being increasingly studied. The biological rationale for the clinical use of PRP includes its effect on the local delivery of growth factors and modification of the inflammatory response and its positive effects on cell proliferation and differentiation4. After activation following intra-articular injection, PRP releases &#945;-granule through degranulation and secretes various growth factors, including the platelet-derived growth factor, the transforming growth factor-&#946;, the insulin-like growth factor, the epidermal growth factor, the vascular endothelial growth factor, and the fibroblast growth factor. These promote osteoblast and chondrocyte proliferation, inhibit cartilage degeneration, strengthen the stability of cartilage and subchondral bone, regulate the gene expression tissue inhibitor of metalloproteinase and maintain the balance of synthesis and degradation of proteoglycans5,6. Therefore, PRP can repair cartilage injury and accelerate bone regeneration.
The outcome of PRP injection is influenced by various factors, including the sampling site7, the type of centrifuge preparation method8, and the use of anticoagulants9 and activators10. There are roughly 3 types centrifugal methods to prepare PRP. Manual centrifugation, equipment-based centrifugation, or plasma filtration techniques are available, although manual and equipment-based methods are the most commonly used. The manual method requires the least equipment, is convenient, is low cost, and is simple to perform (Figure 1). PRP is prepared by performance of manual centrifugation twice. Mixed peripheral blood and anticoagulant are centrifuged to separate hemocytes from plasma and blood platelets. After discarding the red blood cells on the bottom layer, the supernatant liquid is centrifuged for re-separation, dividing it into supernatant platelet-poor plasma, middle PRP, and subnatant residual red blood cells. The middle layer is used for knee joint cavity injection (if the quantity is insufficient, part of the supernatant can be drawn). Evaluations of the method include assessments of blood platelet concentration and clinical outcomes. 
The reporting of PRP preparation protocols in clinical studies is highly inconsistent, and the majority of studies do not provide sufficient information to allow the protocol to be reproduced11. Here, we describe a reproducible method of preparing PRP and treatment of KOA with PRP intra-articular injection, with evaluation of the outcome. Inclusion criteria were patients with knee osteoarthritis who have poor pain relief for simple analgesic medication (such as acetaminophen) and conservative treatment. Exclusion criteria included patients with venous return or lymphatic drainage disorder; patients with knee joint infections; patients with a dermatosis or infection in the injection area; patients with fever; patients with coagulant function abnormality; patients with serious cardiovascular disease. The whole treatment procedure takes less than 30 minutes, and the blood platelet concentration of PRP is proven to reach a standardized measurement. Its effectiveness is demonstrated by evaluating the outcomes during close follow-up.

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	<tbody>
		<tr height="20">
			<td height="20" style="height:20px;width:360px;">Centrifuge</td>
			<td style="width:383px;">Eppendorf</td>
			<td style="width:129px;">5702</td>
			<td style="width:373px;"></td>
		</tr>
		<tr height="20">
			<td height="20" style="height:20px;">Centrifuge tube</td>
			<td>CORNING</td>
			<td>430828</td>
			<td></td>
		</tr>
		<tr height="20">
			<td height="20" style="height:20px;">Horizontal rotor</td>
			<td>Eppendorf</td>
			<td>LL080</td>
			<td></td>
		</tr>
		<tr height="20">
			<td height="20" style="height:20px;">Anerdian III</td>
			<td>Shanghai Likang Disinfectant HiTech Co., LTD</td>
			<td>310173</td>
			<td>Disinfect the skin</td>
		</tr>
		<tr height="20">
			<td height="20" style="height:20px;">1ml Syringe</td>
			<td>KDL&#160; Medical Equipment Co., LTD</td>
			<td>0.4*13 RWLB</td>
			<td></td>
		</tr>
		<tr height="20">
			<td height="20" style="height:20px;">10ml Syringe</td>
			<td>KDL&#160; Medical Equipment Co., LTD</td>
			<td>1.2*38 TWSB</td>
			<td></td>
		</tr>
		<tr height="20">
			<td height="20" style="height:20px;">50ml Syringe</td>
			<td>KDL&#160; Medical Equipment Co., LTD</td>
			<td>0.7*32 TWLB</td>
			<td></td>
		</tr>
		<tr height="20">
			<td height="20" style="height:20px;">Medical Tourniquet</td>
			<td>Changzhou Jinli Latex Products Co., LTD</td>
			<td>0087-2011</td>
			<td></td>
		</tr>
		<tr height="20">
			<td height="20" style="height:20px;">Single-use sterile rubber surgical gloves</td>
			<td>Shanghai jinxiang Latex Products Co., LTD</td>
			<td>17060</td>
			<td></td>
		</tr>
		<tr height="20">
			<td height="20" style="height:20px;">Disposable Draw Blood Needle</td>
			<td>KDL&#160; Medical Equipment Co., LTD</td>
			<td>0.55*20 L(II) RWLB</td>
			<td></td>
		</tr>
		<tr height="20">
			<td height="20" style="height:20px;">Heparin Sodium Injection</td>
			<td>SPH No.1 Biochemical &#38; Pharmaceutical Co., LTD</td>
			<td>1706101</td>
			<td>2ml:12500U</td>
		</tr>
		<tr height="20">
			<td height="20" style="height:20px;">Jifro Hand Antiseptic Rinse Free Gel</td>
			<td>Shanghai Likang Disinfectant HiTech Co., LTD</td>
			<td>311793</td>
			<td></td>
		</tr>
		<tr height="20">
			<td height="20" style="height:20px;">Medical Cotton Swab</td>
			<td>Foshan Kangzheng Medical Supplies Co., LTD</td>
			<td>KZ3-12</td>
			<td>Disinfect the skin</td>
		</tr>
		<tr height="20">
			<td height="20" style="height:20px;">10ml Normal Saline&#160;</td>
			<td>Jiangxi Shuangshi Pharmecutical Co., LTD</td>
			<td>140211458</td>
			<td></td>
		</tr>
		<tr height="20">
			<td height="20" style="height:20px;">Automatic Blood Cell Analyzer</td>
			<td>Beckman Coulter</td>
			<td>LH-750</td>
			<td></td>
		</tr>
		<tr height="20">
			<td height="20" style="height:20px;">Hole-towe</td>
			<td></td>
			<td></td>
			<td>Sterile</td>
		</tr>
		<tr height="20">
			<td height="20" style="height:20px;">Anticoagulation Tube(Blood Collection Tubes, K2E 3.6mg)</td>
			<td>Becton, Dickinson and Company</td>
			<td>CNL17-COO56</td>
			<td>Store in a cool dry place within 4 to 25 degrees Celcius</td>
		</tr>
		<tr height="20">
			<td height="20" style="height:20px;">Tweezers</td>
			<td>RWD LIFE SCIENCE</td>
			<td>F12006-10</td>
			<td></td>
		</tr>
		<tr height="20">
			<td height="20" style="height:20px;">Sterile Gauze</td>
			<td>Guangdong Ze Chang Trade Co., LTD</td>
			<td>170915</td>
			<td></td>
		</tr>
		<tr height="20">
			<td height="20" style="height:20px;">Adhesive Plaster</td>
			<td>3M Transpore</td>
			<td>1527C-0</td>
			<td></td>
		</tr>
		<tr height="20">
			<td height="20" style="height:20px;">Skin Marker Pen</td>
			<td>Guangzhou Mingjia Medical Device Manufacturing Co., LTD</td>
			<td>10110</td>
			<td></td>
		</tr>
	</tbody>
</table>

preparation, procedures and evaluation of platelet-rich plasma injection in the treatment of knee osteoarthritis

Knee osteoarthritis (KOA) is one of the most frequently encountered diseases in the orthopedic department. Existing non-surgical treatments have a limited effect on the repair of cartilage and on bone regeneration. Platelet-rich plasma (PRP) is an autologous bioactive substance that can repair cartilage injury and accelerate bone regeneration effectively. However, reporting of PRP preparation protocols in clinical studies is highly inconsistent, with the majority of studies providing insufficient information to allow the protocol to be reproduced. We describe a repeatable method of preparing PRP visually, the treatment of KOA using PRP intra-articular injection, and methods of evaluating the outcome. PRP was prepared using manual double centrifugation. The PRP layer was extracted from peripheral blood and used for knee joint cavity injection. Evaluations included assessments of blood platelet concentrations and clinical outcomes. Preparation of PRP by manual centrifugation requires less apparatus and is less costly than plasma filtration or centrifugation using equipment. The centrifugation time of our double centrifugation method was 6 and 5 minutes for the respective centrifugations at forces of 800 and 1400 x g, respectively, to allow for the consistent preparation of standardized PRP. However, a manual method is susceptible to operator error, and PRP batch preparation is not available. Intra-articular injection of PRP proved to be an effective treatment for knee osteoarthritis. The entire treatment procedure took less than 30 minutes, the blood platelet concentration of PRP could be standardized, and treatment was proven to be effective when evaluated by follow-up.

As a result, the platelet count of the PRP reached a standard concentration level of 1121 x 103/&#181;L. We conducted the 15 follow-up surveys described in the protocol on a 55-year-old male patient with early KOA. It was obvious that early clinical outcome was satisfactory after the intra-articular administration of PRP (Figure 2). However, medium-term efficacy was slightly inferior. A markedly significant analgesic effect was observed (<strong cl...

Watch this Scientific Journal Video about Preparation, Procedures and Evaluation of Platelet-Rich Plasma Injection in the Treatment of Knee Osteoarthritis at JoVE.com

Preparation, Procedures and Evaluation of Platelet-Rich Plasma Injection in the Treatment of Knee Osteoarthritis

Knee osteoarthritis is frequently seen in the orthopedic department. We introduce in detail the entire knee osteoarthritis treatment process with platelet-rich plasma injection, including preparation, procedures, and evaluation.

Knee osteoarthritis (KOA) is one of the most frequently encountered diseases in the orthopedic department. Existing non-surgical treatments have a limited ...

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