This work was supported by Shanghai Critical Clinical Specialties Construction Project (Grant Number: Shslczdzk04001); the Sailing program of Shanghai Science and Technology Commission (Grant Number: 22YF1444300); Projects within the budget of Shanghai University of Traditional Chinese Medicine (Grant Number: 2021LK091).

Standardized Tuina Manipulation for Knee Osteoarthritis in Rats

<ol>
	<li>Joint Surgery Group of Chinese Orthopaedic Association, Chinese Association of Orthopaedic Surgeons, National Clinical Research Center for Geriatric Diseases, Chinese Journal of Orthopaedics. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Chinese+Osteoarthritis+Treatment+Guidelines+(2021+Edition).">Chinese Osteoarthritis Treatment Guidelines (2021 Edition).</a> Chinese Journal of Orthopaedics. 41 (18), 24 (2021).</li><li>David, S., 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=Epidemiology+of+knee+osteoarthritis+in+general+practice:+a+registry-based+study.">Epidemiology of knee osteoarthritis in general practice: a registry-based study.</a> BMJ Open. 10 (1), 031734 (2020).</li><li>Callahan, L. F., Cleveland, R. J., Allen, K. 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The authors declare that they have no competing interests.

This study provides a protocol for Tuina manipulation on KOA rats. Through pain behavior tests and histomorphological findings, it suggested that such a series of Tuina manipulation applied to KOA rats could reduce synovial inflammation and cartilage apoptosis, which could be a reference of Tuina manipulation on animal models of KOA.
There are several critical procedures during the protocol. First, it&#39;s important to choose an appropriate method for inducing the KOA model. There are various methods to induce KOA model, including joint cavity injection25, intra-articular surgery26,27, joint braking method28,29, etc. Since KOA models induced by invasive methods would leave wounds near the joints, which may interfere with the effect of Tuina manipulation, we chose the non-invasive method of joint immobilization. The joint immobilization method can be divided into hyper-extension and hyper-flexion fixation. Shang et al. compared the effects of these two fixation methods and found there was little difference in the effect of cartilage apoptosis23. However, their experimental subjects were rabbits, which are larger in size and relatively easy to fix compared to rats. Fixed plaster in a flexion position is more likely to fall off under rats gnawing. Therefore, we chose the immobilization method of hyper-extension fixation to induce KOA. We referred to He et al. and used the plaster bandage with a tongue depressor as the fixation device30. However, the gnawing ability of rats was stronger than expected, and the device could not serve as a fixation. Later we found a kind of denture base material, which can form a moldable but hard shell on the outside of the plaster, and effectively reduce the wear and tear of the fixation device by rats gnawing.&#160;A tight fixation would affect the blood circulation of the lower limbs, while too loose fixation would be easy to fall off. Therefore, the circulation of the lower limbs of the rats should be observed daily during the 3 weeks of modeling. The fixation should be released when the lower limbs become swollen and purplish. Re-install the fixation when it is loose, and the rats can flex their knees. During this study, the rats were not restricted from socializing during modeling, but it was found that they would chew each other&#39;s fixation to help each other break free. Perhaps isolating the rats would have resulted in better modeling. However, isolation might contribute to the rats&#39; depression and stereotypical behavior.&#160;
We consider that the key point of Tuina manipulation is to keep the consistency of the intensity and frequency. Previous research concluded that the optimal intensity for animal Tuina should be 80% of its maximum tolerable intensity31. At this point, the rats should show signs of receiving mechanical stimulation, but with no signs of pain or paw retraction. We tested the maximum tolerable intensity of rats, which is around 5-8 N. So, we set the pushing intensity to 3-5 N, which may lead to better efficacy. The frequency of the manipulation is 2 Hz, according to the textbook of Tuina. Previous studies have not set a standardization for Tuina manipulation, and the intensity and frequency might vary after operating different acupoints and altering the left and right hands during the experiment, which can affect the accuracy of the results. The finger pressure recordings used in this experiment provide real-time observation of the intensity and frequency of the manipulation during the experiment, which can keep the consistency of the manipulation. Some scholars use machines to simulate Tuina manipulation and apply it on animals, which has the advantage of standardization of the manipulation32.
For the selection of acupoints, we referred to the study of Wang and Liu and chose five acupoints around the knee joint to facilitate manipulation33,34. We chose the plaster-immobilized method to induce KOA, which mostly ends up with muscle atrophy and joint stiffness35. ST34 and ST35 belong to the foot Yangming stomach meridian which is critical in treating atrophy (mostly manifested as weakness and muscle atrophy) in traditional Chinese medicine theory and has good adaptability for KOA induced by the plaster-immobilized method. EX-LE4 is a common acupoint for the clinical treatment of KOA, mostly used together with ST35. Tan et al. found that acupuncture and moxibustion on ST35, ST36, and EX-LE4 could reduce inflammatory factor expression in the synovial membrane36. SP10 and BL40 are the core acupoints of clinical Tuina treatment for KOA37,38, and SP10 is most frequently used in Tuina prescriptions for KOA39.
A difficult part of Tuina manipulation is the fixation of rats. Rats may struggle or avoid manipulation, which increases the difficulty of performing the experiment. Although rat immobilizers can immobilize rats and expose their hind limbs, the development of KOA limits hind limb flexion and extension. Therefore, settling rats with immobilizers may lead to pain and incomplete exposure of the hind limbs, which may affect Tuina manipulation. We designed a single-exit cloth bag based on the rat&#39;s preference for dark environments and its tendency to burrow. Close the cloth bag when the rat enters, which is, the head and forelimbs are located in the bag, and its hind limbs are exposed outside. This method can effectively reduce the struggle of rats and keep them quiet during Tuina.
The results of the pre-experiment suggest that Tuina manipulation may reduce the inflammatory response of synovial tissue and decrease the apoptosis of chondrocytes, thus acting as a treatment for KOA. However, more studies are needed to verify this.
There are several shortcomings of this protocol. First, the small size of rats makes it difficult to locate the acupoints precisely. The operator&#39;s fingers are too large compared to the acupoints on rats, which may affect the effect of Tuina manipulation. We had considered adding a rubber particle of about 2 mm in diameter on the fingertip to increase the accuracy of stimulating acupoints. But the particles might lead to higher intensity of pressure and increase the difficulty in controlling the consistency. In addition, it was more likely to damage the finger pressure recordings, so we did not choose this method. Second, our study ignored the change of inflammatory factors in the synovial fluid and chondrocyte apoptosis, which may weaken the credibility of the study. We will intensify research in this area in the future. Third, press and kneading manipulation is one of the Tuina manipulations, and cannot encompass the effects of Tuina in treating KOA. Other Tuina manipulations implemented in animals need to be further explored, including joint movement manipulations, and rubbing manipulation. Moreover, positive control would better show the effectiveness of the manipulation, but it was not designed in this study, and we will add this in a follow-up study.
Overall, this study provides a standardized protocol for Tuina manipulation on KOA rats, and the effectiveness of the manipulation has been verified by pain behavior tests and microscopical findings. The protocol preliminary proves that the therapeutic effects of Tuina may be related to reducing synovial inflammation and delayed chondrocyte apoptosis. More research is needed to explore the mechanism of Tuina for KOA.

Knee osteoarthritis (KOA) is a degenerative disease mainly manifested in joint pain. Fibrosis, cracking, ulceration, and loss of articular cartilage are the main causes of this disease1. KOA has a high prevalence and may result in a profound impact on the daily life of patients, causing disability in severe cases. Among people aged 45-84 years, the prevalence of KOA increases with age, and the prevalence among people aged 85 years and above is 15%, with a predominance in women2,3. In addition, KOA might bring a serious economic burden on both the individual and the society. A survey showed that direct health care costs on KOA per capita reached up to $8,858 &#177; $5,120 per year4. With the aging of society, KOA has become a worldwide health problem and a major social issue, as well as a topical issue for scientific research.
Evidence-based studies have shown the effectiveness of Tuina manipulation in treating KOA5. Tuina manipulation could relieve pain and improve dysfunction in KOA patients, the mechanism of which is related to anti-inflammatory effects6,7. Scholars found that Tuina manipulation effectively inhibited the expression of inflammatory factors interleukin (IL)-&#946; and 5-hydroxytryptamine and slowed down the degeneration of articular cartilage in a rabbit KOA model8. It suggested that Tuina could promote blood circulation and metabolism at the lesion site, which helped clear inflammatory factors such as IL-1, IL-6 and tumor necrosis factor (TNF)-&#945;, thereby alleviating the clinical symptoms of KOA9. In addition, the passive movement of the joint through Tuina manipulation can promote the penetration and diffusion of synovial fluid into the articular cartilage and improves tissue nutrient metabolism10. Other studies suggested that Tuina manipulation can effectively improve the biomechanical indexes in KOA patients11. Manipulations applied on soft tissues can improve stress distribution over limbs and enhance balance function12,13. At the same time, with some joint adjustment manipulations, the alignment of the lower limbs can also be adjusted to correct abnormal gaits14,15.
The mechanism of action of Tuina manipulation in treating KOA remains to be explored, and therefore, an experimental study is necessary. The key to the application of Tuina in experimental animals is the standardization of modeling, animal fixation and intervention methods16. The modeling method determines whether the experimental animal can exhibit the characteristics of the disease. Meanwhile, appropriate fixation methods can facilitate the intervention of the Tuina manipulation and better reflect the effect of Tuina. The standardization of intervention methods is the most difficult part of Tuina manipulation. In 2010, the basic system of Chinese acupuncture standards mentioned the acupuncture point standards for experimental animals, providing the possibility for acupuncture and Tuina operations in animal experiments17. However, there are still difficulties in standardizing Tuina manipulation. There are multiple types of Tuina manipulation18. The choice of the specific manipulation mainly depends on the disease to be treated and the therapeutic theories the performer prefers. In the study of Tuina for KOA, more attention has been paid to the point-pressing manipulation (pressing the specific acupoints with the thumb or elbow), Yizhichan pushing manipulation (a pushing manipulation by wiggling the thumb), and press and kneading manipulation (which refers to pressing and kneading the specific area of the body surface by finger or palm)19. Press and kneading manipulation is one of the most widely used Tuina manipulations, which combines pressing and kneading to move the subcutaneous tissue20. Press and kneading manipulation applied on acupoints can promote blood circulation and relieve pain and represents the therapeutic effect of Tuina on KOA19.
In this protocol, the operation of press and kneading manipulation on KOA rats will be described in detail, including selected acupoints, intensity and frequency of the manipulation and the body position of the rat, so as to provide a reference for future research.

<table><tbody><tr><td>absolute ethanol</td><td>Supelco</td><td>PHR1070</td><td>For making specimen</td></tr><tr><td>ALMEMO admeasuring apparatus</td><td>ahlborn</td><td>2450-1</td><td>For Mechanical Withdrawal Threshold test</td></tr><tr><td>Anti-Digoxin antibody</td><td>Sigma-Aldrich</td><td>SAB4200669</td><td>For HE stain IHC or TUNEL</td></tr><tr><td>Anti-IL-1 beta</td><td>abcam</td><td>ab283818</td><td>For HE stain IHC or TUNEL</td></tr><tr><td>DAB Substrate kit</td><td>Solarbio</td><td>DA1010</td><td>For HE stain IHC or TUNEL</td></tr><tr><td>Denture base materials</td><td>Shanghai New Century</td><td>20000356</td><td>For model making</td></tr><tr><td>eosin</td><td>bioswamp&amp;nbsp;</td><td>PAB180016&amp;nbsp;</td><td>For HE stain IHC or TUNEL</td></tr><tr><td>Finger pressure recordings</td><td>Suzhou Changxian Optoelectronic Technology</td><td>CX1003w</td><td>For Tuina manipulation</td></tr><tr><td>formic acid solution</td><td>Sigma-Aldrich</td><td>695076</td><td>For decalcification</td></tr><tr><td>H2O2</td><td>Sigma-Aldrich</td><td>386790-M</td><td>For HE stain IHC or TUNEL</td></tr><tr><td>hematoxylin</td><td>bioswamp&amp;nbsp;</td><td>PAB180015</td><td>For HE stain IHC or TUNEL</td></tr><tr><td>Isoflurane</td><td>Shanghai Yuyan Scientific Instrument Company</td><td>S10010533</td><td>For gas anesthesia</td></tr><tr><td>neutral resins</td><td>bioswamp&amp;nbsp;</td><td>PAB180017</td><td>For HE stain IHC or TUNEL</td></tr><tr><td>Paraformaldehyde Fix Solution</td><td>Sigma-Aldrich</td><td>100496</td><td>For histology</td></tr><tr><td>PBS</td><td>Sigma-Aldrich</td><td>P3813</td><td>For HE stain IHC or TUNEL</td></tr><tr><td>Plantar Test Apparatus</td><td>IITC Life Science</td><td>/</td><td>For Paw Withdrawal Latency test</td></tr><tr><td>plaster of Paris bandage</td><td>WANDE</td><td>20150023</td><td>For model making</td></tr><tr><td>Proteinase K</td><td>Sigma-Aldrich</td><td>124568</td><td>For HE stain IHC or TUNEL</td></tr><tr><td>TNF Alpha Monoclonal antibody</td><td>Proteintech</td><td>60291-1-Ig</td><td>For HE stain IHC or TUNEL</td></tr><tr><td>TUNEL&amp;nbsp;</td><td>Servicebio</td><td>GDP1042</td><td>For HE stain IHC or TUNEL</td></tr><tr><td>Wax</td><td>Sigma-Aldrich</td><td>327204</td><td>For making specimen</td></tr><tr><td>xylene</td><td>Shanghai Sinopharm Group</td><td>100092</td><td>For making specimen</td></tr></tbody></table>

tuina manipulation to reduce inflammation and cartilage loss in knee osteoarthritis rats

Clinical trials suggest that Tuina manipulation is effective in treating knee osteoarthritis (KOA), while further studies are required to discover its mechanism. Therefore, the manipulation of animal models of knee osteoarthritis is critical. This protocol provides a standard process for Tuina manipulation on KOA rats and a preliminary exploration of the mechanism of Tuina for KOA. The press and kneading manipulation method (a kind of Tuina manipulation that refers to pressing and kneading the specific area of the body surface) is applied on 5 acupoints around the knee joint of rats. The force and frequency of the manipulation were standardized by finger pressure recordings, and the position of the rat during manipulation is described in detail in the protocol. The effect of manipulation can be measured by pain behavior tests and microscopic findings in synovial and cartilage. KOA rats showed significant improvement in pain behavior. The synovial tissue inflammatory infiltration was reduced in the Tuina group, and the expression of tumor necrosis factor (TNF)-&#945; was significantly lower. Compared to the control group, chondrocyte apoptosis was less in the Tuina group. This study provides a standardized protocol for Tuina manipulation on KOA rats and preliminary proof that the therapeutic effects of Tuina may be related to reducing synovial inflammation and delayed chondrocyte apoptosis.

Pain behavior tests 
The MWT results showed that the MWT of the right hind limb after modeling was significantly lower than before (p&#60;0.05). Compared with the control group, the MWT of rats was significantly elevated after Tuina (p&#60;0.05; Figure 5 and Table 2).
<img alt="Figure 5" class="xfigimg" src="/files/ftp_upload/65451/65451fig05.jpg" /> 
Figure 5. Results of the mechanical withdrawal threshold test (<img alt="Equation 1" src="/files/ftp_upload/65451/65451eq01.jpg" style="margin: auto;" />, Newton). There were 5 rats each in the Tuina group and the control group. The MWT of the rats at different time points is shown in the figure. After the modeling, the MWT of the rats decreased significantly, suggesting that the pain of the rats was aggravated, and the KOA model was successfully prepared. Subsequently, the MWT gradually improved, suggesting pain relief. The generalized estimation equation was used for statistical calculation. The difference in MWT between the Tuina group and the control group was statistically significant at day 21 compared to that after modeling. The comparison between the two groups was statistically significant at D7, D14 and D21, *p&#60;0.05. Moreover, rats in Tuina group have higher MWT than that in the control group. <a href="https://www-jove-com.remotexs.ntu.edu.sg/files/ftp_upload/65451/65451fig05large.jpg" target="_blank">Please click here to view a larger version of this figure.</a>
Table 2. Mechanical Withdrawal Threshold ( <img alt="Equation 1" src="/files/ftp_upload/65451/65451eq01.jpg" style="margin: auto;" />, N). Comparison before and after modeling, #p&#60;0.05. Comparison between Tuina group and control group, *p&#60;0.05. <a href="https://www-jove-com.remotexs.ntu.edu.sg/files/ftp_upload/65451/Table.2_Re.xlsx" target="_blank">Please click here to download this Table.</a>
The PWL results showed that the PWL of the right hind limb after modeling was significantly shorter than before (p&#60;0.05). Compared with the control group, the MWT of rats was significantly prolonged after Tuina (p&#60;0.001; Figure 6 and Table 3).
<img alt="Figure 6" class="xfigimg" src="/files/ftp_upload/65451/65451fig06.jpg" /> 
Figure 6. Results of the paw withdrawal latency test (<img alt="Equation 1" src="/files/ftp_upload/65451/65451eq01.jpg" style="margin: auto;" />, second). There were 5 rats each in the Tuina group and the control group. The PWL of the rats at different time points is shown in the figure. After the modeling, the PWL of the rats decreased significantly, suggesting that the pain of the rats was aggravated, and the KOA model was successfully prepared. At first, the improvement of PWT in the Tuina group was slower than that in the control group. After D7, rats in the Tuina group improved rapidly and surpassed the control group at D21. The generalized estimation equation was used for statistical calculation. The difference in MWT between the Tuina group and the control group was statistically significant at day 21 compared to that after modeling. The comparison between the two groups was statistically significant at D7, D14 and D21, ***p&#60;0.001. <a href="https://www-jove-com.remotexs.ntu.edu.sg/files/ftp_upload/65451/65451fig06large.jpg" target="_blank">Please click here to view a larger version of this figure.</a>
Table 3. Paw Withdrawal Latency (<img alt="Equation 1" src="/files/ftp_upload/65451/65451eq01.jpg" style="margin: auto;" />, s). Comparison before and after modeling, #p&#60;0.05. Comparison between the Tuina group and the control group, ***p&#60;0.001. <a href="https://www-jove-com.remotexs.ntu.edu.sg/files/ftp_upload/65451/Table.3_RE.xlsx" target="_blank">Please click here to download this Table.</a>
Histomorphological assay 
Upon analyzing the synovial membrane in the control group, inflammatory cell infiltration and fibrous tissue hyperplasia were seen in synovial tissue. The synovial cells were disorganized, and a small amount of capillary hyperplasia was seen around the synovial tissue (Figure 7).
<img alt="Figure 7" class="xfigimg" src="/files/ftp_upload/65451/65451fig07.jpg" /> 
Figure 7. Microscopic observation of the synovial tissue. (A) Synovial tissue in the control group. The disorganized arrangement of synovial cells is seen in the figure. The proliferating vessels are marked with red arrows, and inflammatory cells are marked with black arrows. (B) Synovial tissue in Tuina group. Synovial cells were more neatly arranged. Inflammatory cells are mainly distributed at the edges rather than infiltrating inwards. The proliferating vessels are marked with red arrows, and inflammatory cells are marked with black arrows. <a href="https://www-jove-com.remotexs.ntu.edu.sg/files/ftp_upload/65451/65451fig07large.jpg" target="_blank">Please click here to view a larger version of this figure.</a>
In the Tuina group, synovial cells were neatly arranged, with a small amount of inflammatory cell infiltration, fibrous tissue hyperplasia, and a small amount of capillary hyperplasia visible at the tissue margins.
Upon analyzing the cartilage, it was seen that the TUNEL staining positive area in the Tuina group was significantly smaller than that in the control group, indicating that there were fewer apoptotic chondrocytes in the Tuina group (Figure 8).
<img alt="Figure 8" class="xfigimg" src="/files/ftp_upload/65451/65451fig08.jpg" /> 
Figure 8.&#160;TUNEL staining of the cartilage. (A) Cartilage in the control group. The red part of the figure is the positive area of TUNEL staining. (B) Cartilage in Tuina group. The red part of the figure is the positive area of TUNEL staining. The positive area in the Tuina group is significantly smaller than that in the control group. <a href="https://www-jove-com.remotexs.ntu.edu.sg/files/ftp_upload/65451/65451fig08large.jpg" target="_blank">Please click here to view a larger version of this figure.</a>
Immunohistochemistry 
For TNF-&#945;, there was a significant difference in synovial TNF-&#945; expression between the two groups, and the expression in the Tuina group was significantly lower than that in the control group (Table 4).
Table 4. TNF-&#945; Expression in Synovium (<img alt="Equation 1" src="/files/ftp_upload/65451/65451eq01.jpg" style="margin: auto;" />, *10-2). Two independent sample t-test was used for statistical analysis, *p&#60;0.05. <a href="https://www-jove-com.remotexs.ntu.edu.sg/files/ftp_upload/65451/Table 4_RE.xlsx" target="_blank">Please click here to download this Table.</a>
For IL-1&#946;, there was no significant difference in the amount of synovial IL-1&#946; expression between the two groups, as measured by the statistics of Image J. However, the mean value of the Tuina group was subtly lower, indicating less expression (Table 5).
Table 5. IL-&#946; Expression in Synovium (<img alt="Equation 1" src="/files/ftp_upload/65451/65451eq01.jpg" style="margin: auto;" />, *10-2). Two independent sample t-test was used for statistical analysis. <a href="https://www-jove-com.remotexs.ntu.edu.sg/files/ftp_upload/65451/Table 5_RE.xlsx" target="_blank">Please click here to download this Table.</a>

Watch this Scientific Journal Video about Understanding the Mechanism of Tuina Manipulation in Rats at JoVE.com

Author Spotlight: Understanding the Mechanism of Tuina Manipulation in Rats 

We present a protocol for Tuina manipulation performed on plaster-immobilized-induced knee osteoarthritis rats. Based on the preliminary results, it suggested that the efficacy of the method relied on the reduction of inflammation and cartilage loss.

Tuina Manipulation to Reduce Inflammation and Cartilage Loss in Knee Osteoarthritis Rats

Clinical trials suggest that Tuina manipulation is effective in treating knee osteoarthritis (KOA), while further studies are required to discover its ...

tuina-manipulation-to-reduce-inflammation-cartilage-loss-knee

Nanyang Technological University

Research

JoVE Journal

Medicine

1.9K Views.  Yueyang Hospital of Integrated Traditional Chinese and Western Medicine Affiliated to Shanghai University of Traditional Chinese Medicine. We bestudeerden het mechanisme van Tuina-manipulatie bij de behandeling van knieartrose. We ontdekten dat Tuina effectief is voor pijnstillende en verbeterende motorische functies. Er zijn echter nog steeds standaardprotocollen voor Tuina of KOA en de conclusies zijn niet betrouwbaar genoeg, dus we proberen een standaardprotocol te bouwen.Onlangs zijn veel teams begonnen met het ontwikkelen van Tuina op massagerobots, die speciale mechanische golfvormen kunnen uitvoeren voor manipulati...

Video: Auteur in de schijnwerpers: het mechanisme van Tuina-manipulatie bij ratten begrijpen 

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.

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 ...

Using a Knee Arthrometer to Evaluate Tissue-specific Contributions to Knee Flexion Contracture in the Rat

Normal knee range of motion (ROM) is critical to well-being and allows one to perform basic activities such as walking, climbing stairs and sitting. Lost ROM is called a joint contracture and results in increased morbidity. Due to the difficulty of reversing established knee contractures, early detection is important, and hence, knowing risk factors for their development is essential. The rat represents a good model with which the effect of an intervention can be studied due to the similarity of rat knee anatomy to that of humans, the rat's ability to tolerate long durations of knee immobilization in flexion, and because mechanical data can be correlated with histologic and biochemical analysis of knee tissue.
Using an automated arthrometer, we demonstrate a validated, precise, reproducible, user-independent method of measuring the extension ROM of the rat knee joint at specific torques. This arthrometer can be used to determine the effects of interventions on knee joint ROM in the rat.

The goal of the protocol is to measure the extension range of motion of the rat knee. The effects of various diseases that increase the stiffness of the knee joint and the effectiveness of treatments can be quantified.

Normal knee range of motion (ROM) is critical to well-being and allows one to perform basic activities such as walking, climbing stairs and sitting. Lost ...

The Lower Body Positive Pressure Treadmill for Knee Osteoarthritis Rehabilitation

Here, based on a clinician&rsquo;s point-of-view, we propose a two-model lower body positive pressure (LBPP) protocol (walking and squatting models) in addition to a clinical, functional assessment methodology, including details for further encouragement of the development of non-drug surgical intervention strategies in knee osteoarthritis patients. However, we only present the effect of LBPP training in improvement of pain and knee function in one patient through three-dimensional gait analysis. The exact, long-term effects of this approach should be explored in future studies.

Here, based on a clinician&#8217;s point-of-view, we propose a two-model lower body positive pressure (LBPP) protocol (walking and squatting models) in addition to a clinical, functional assessment methodology, including details for further encouragement of the development of non-drug surgical intervention strategies in knee osteoarthritis patients. However, we only present the effect of LBPP training in improvement of pain and knee function in one patient through three-dimensional gait analysis. The exact, long-term effects of this approach should be explored in future studies.

Here, based on a clinician&rsquo;s point-of-view, we propose a two-model lower body positive pressure (LBPP) protocol (walking and squatting models) in ...

Three-Dimensional Finger Motion Tracking during Needling: A Solution for the Kinematic Analysis of Acupuncture Manipulation

Three-dimensional (3D) motion tracking has been used in many fields, such as the researches of sport and medical skills. This experiment aimed to use 3D motion tracking technology to measure the kinematic parameters of the joints of fingers during acupuncture manipulation (AM) and establish three technical indicators "amplitude, velocity and time". This method can reflect the operation characteristics of AM and provide quantitative parameters along three axes of multiple finger joints. The current evidence shows that the method has great potential for future applications such as the study of the dose-effect relationship of acupuncture, teaching, and learning of AM, and the measurement and preservation of famous acupuncturists' AM.

This experimental method describes a solution for the kinematic analysis of acupuncture manipulation with three-dimensional finger motion tracking technology.

Three-dimensional (3D) motion tracking has been used in many fields, such as the researches of sport and medical skills. This experiment aimed to use 3D ...

Destabilization of the Medial Meniscus and Cartilage Scratch Murine Model of Accelerated Osteoarthritis

Osteoarthritis is the most prevalent musculoskeletal disease in people over 45, leading to an increasing economic and societal cost. Animal models are used to mimic many aspects of the disease. The present protocol describes the destabilization and cartilage scratch model (DCS) of post-traumatic osteoarthritis. Based on the widely used destabilization of the medial meniscus (DMM) model, DCS introduces three scratches on the cartilage surface. The current article highlights the steps to destabilize the knee by transecting the medial meniscotibial ligament followed by three intentional superficial scratches on the articular cartilage. The possible analysis methods by dynamic weight-bearing, microcomputed tomography, and histology are also demonstrated. While the DCS model is not recommended for studies that focus on the effect of osteoarthritis on the cartilage, it enables the study of osteoarthritis development in a shorter time window, with special focus on (1) osteophyte formation, (2) osteoarthritic and injury pain, and (3) the effect of cartilage damage in the whole joint.

The present protocol describes the controlled microblade scratches on the surface of the articular cartilage after destabilizing the mouse knee by cutting the medial miniscotibial ligament. This animal model presents an accelerated form of osteoarthritis (OA) suitable for studying osteophyte formation, osteosclerosis, and early-stage pain.

Osteoarthritis is the most prevalent musculoskeletal disease in people over 45, leading to an increasing economic and societal cost. Animal models are ...

FSN Therapy for Knee Osteoarthritis Pain Management

Fu's Subcutaneous Needling for Knee Osteoarthritis Pain

Fu's subcutaneous needling (FSN) is a new acupuncture and dry needling technique based on traditional Chinese medicine. It rapidly produces long-lasting effects in soft tissue injuries, particularly in painful musculoskeletal conditions, by providing stimulation primarily in the subcutaneous area. Osteoarthritis (OA) is the most common joint disease in adults worldwide and is often accompanied by a painful syndrome of structural changes in the peripheral joints of the knee. However, the etiology of OA pain is not fully understood, though myofascial trigger points (MTrPs) are commonly found in the lower limb muscles (so-called "tightened muscles") of patients with knee OA. 
FSN has been used in many fields for the treatment of acute pain problems and can relieve muscle contraction from MTrPs, thereby improving the local circulation. This study recruited patients with pain from knee OA into an FSN group or a transcutaneous electrical nerve stimulation (TENS) group with three treatment sessions and a follow-up over the course of 2 weeks. The results showed that FSN was effective in treating soft tissue pain around the knee with OA. This study aimed to establish and visualize three key technical indicators during FSN therapy, including the FSN needle insertion point and layer; the frequency and duration of the swaying movement; and the manipulation of the reperfusion approach. These findings have great potential for future applications in myofascial pain treatment, especially for pain management. Following this protocol could enhance FSN skills.

Author Spotlight: Fu's Subcutaneous Needling for Knee Osteoarthritis Pain  

We present a protocol for using Fu's subcutaneous needling for knee osteoarthritis pain, which combines swaying movement and reperfusion approach techniques. This protocol has great potential for future applications in myofascial pain treatment and could enhance Fu's subcutaneous needling (FSN) manipulation skills.

Fu's subcutaneous needling (FSN) is a new acupuncture and dry needling technique based on traditional Chinese medicine. It rapidly produces long-lasting ...

Tuina Therapy for Frozen Shoulders in a Rat Model

Tuina in a Frozen Shoulder Rat Model: An Efficient and Reproducible Protocol

Frozen shoulder (FS) is a common condition with no defined optimal therapy. Tuina therapy, a traditional Chinese medicine (TCM) technique used to treat FS patients in Chinese hospitals, has demonstrated excellent results, but its mechanisms are not fully understood. Building on a previous study, this work aimed to develop a Tuina protocol for an FS rat model. We randomly divided 20 SD rats into control (C; n = 5), FS model (M; n = 5), FS model Tuina treatment (MT; n = 5), and FS model oral treatment (MO; n = 5) groups. This study used the cast immobilization method to establish the FS rat model. The effect of Tuina and oral dexamethasone on the glenohumeral range of motion (ROM) was evaluated, and the histological findings were assessed. Our study showed that Tuina and oral dexamethasone were able to improve shoulder active ROM and preserve the structure of the capsule, with Tuina therapy proving to be more effective than oral dexamethasone. In conclusion, the Tuina protocol established in this study was highly effective for FS.

Author Spotlight: Investigating the Effectiveness of Tuina Therapy for Frozen Shoulder 

This study develops an efficient and reproducible Tuina protocol for treating frozen shoulder established in a rat model. This approach will help to study the Tuina therapy treatment method for frozen shoulders.

Frozen shoulder (FS) is a common condition with no defined optimal therapy. Tuina therapy, a traditional Chinese medicine (TCM) technique used to treat FS ...

Treating Knee Osteoarthritis with Tuina in a Rabbit Model

Tuina Intervention in Rabbit Model of Knee Osteoarthritis

Knee osteoarthritis (KOA) is mainly characterized by degenerative changes in the knee joint's cartilage and surrounding soft tissues. The efficacy of Tuina in treating KOA has been confirmed, but the underlying mechanism needs to be investigated. This study aims to establish a scientifically feasible KOA rabbit model treated with Tuina to reveal the underlying mechanisms. For this, 18, 6-month-old normal-grade male New Zealand rabbits were randomly divided into sham, model, and Tuina groups, with 6 rabbits in each group. The KOA model was established by injecting 4% papain solution into the knee joint cavity. The Tuina group was intervened with Tuina combined with the knee joint rotary correction method for 4 weeks. Only the standard grasping and fixation were performed in sham and model groups. At the end of the 1-week intervention, the knee joint range of motion (ROM) was observed, and cartilage hematoxylin-eosin (HE) staining was done. The study shows that Tuina could inhibit chondrocyte apoptosis, repair cartilage tissue, and restore knee joint ROM. In conclusion, this study demonstrates the scientific feasibility of Tuina treatment for KOA model rabbits, highlighting its potential application in the study of KOA and similar knee joint-related conditions.

Author Spotlight: Using a Rabbit Model to Explore the Efficacy of Tuina in Treating Knee Osteoarthritis 

The protocol describes a method for Tuina intervention in a rabbit model of knee osteoarthritis.

Knee osteoarthritis (KOA) is mainly characterized by degenerative changes in the knee joint's cartilage and surrounding soft tissues. The efficacy of ...

A Reproducible Cartilage Impact Model to Generate Post-Traumatic Osteoarthritis in the Rabbit

Post-traumatic osteoarthritis (PTOA) is responsible for 12% of all osteoarthritis cases in the United States. PTOA can be initiated by a single traumatic event, such as a high-impact load acting on articular cartilage, or by joint instability, as occurs with anterior cruciate ligament rupture. There are no effective therapeutics to prevent PTOA currently. Developing a reliable animal model of PTOA is necessary to better understand the mechanisms by which cartilage damage proceeds and to investigate novel treatment strategies to alleviate or prevent the progression of PTOA. This protocol describes an open, drop tower-based rabbit femoral condyle impact model to induce cartilage damage. This model delivered peak loads of 579.1 &plusmn; 71.1 N, and peak stresses of 81.9 &plusmn; 10.1 MPa with a time-to-peak load of 2.4 &plusmn; 0.5 ms. Articular cartilage from impacted medial femoral condyles (MFCs) had higher rates of apoptotic cells (p = 0.0058) and possessed higher Osteoarthritis Research Society International (OARSI) scores of 3.38 &plusmn; 1.43 compared to the non-impacted contralateral MFCs (0.56 &plusmn; 0.42), and other cartilage surfaces of the impacted knee (p &lt; 0.0001). No differences in OARSI scores were detected among the non-impacted articular surfaces (p &gt; 0.05).

The open medial femoral condyle impact model in rabbits is reliable for studying post-traumatic osteoarthritis (PTOA) and novel therapeutic strategies to mitigate PTOA progression. This protocol generates an isolated cartilage defect of the posterior medial femoral condyle in rabbits using a carriage-based drop tower with an impactor head.

Post-traumatic osteoarthritis (PTOA) is responsible for 12% of all osteoarthritis cases in the United States. PTOA can be initiated by a single traumatic ...

Therapeutic Potential of Tuina Massage for Knee Osteoarthritis

Tuina Intervention in Sodium Monoiodoacetate Injection-Induced Rat Model of Knee Osteoarthritis

Knee osteoarthritis (KOA), a common degenerative joint disorder, is characterized by chronic pain and disability, which can progress to irreparable structural damage of the joint. Investigations into the link between articular cartilage, muscles, synovium, and other tissues surrounding the knee joint in KOA are of great importance. Currently, managing KOA includes lifestyle modifications, exercise, medication, and surgical interventions; however, the elucidation of the intricate mechanisms underlying KOA-related pain is still lacking. Consequently, KOA pain remains a key clinical challenge and a therapeutic priority. Tuina has been found to have a regulatory effect on the motor, immune, and endocrine systems, prompting the exploration of whether Tuina could alleviate KOA symptoms, caused by the upregulation of inflammatory factors, and further, if the inflammatory factors in skeletal muscle can augment the progression of KOA.
We randomized 32 male Sprague Dawley (SD) rats (180-220 g) into four groups of eight animals each: antiPD-L1+Tuina (group A), model (group B), Tuina (group C), and sham surgery (group D). For groups A, B, and C, we injected 25 &#181;L of sodium monoiodoacetate (MIA) solution (4 mg MIA diluted in 25 &#181;L of sterile saline solution) into the right knee joint cavity, and for group D, the same amount of sterile physiological saline was injected. All the groups were evaluated using the least to most stressful tests (paw mechanical withdrawal threshold, paw withdrawal thermal latency, swelling of the right knee joint, Lequesne MG score, skin temperature) before injection and 2, 9, and 16 days after injection.

Author Spotlight: Treating Knee Osteoarthritis with Tuina - A New Perspective 

This protocol describes the methods of Tuina intervention in sodium monoiodoacetate injection-induced rat model of knee osteoarthritis (KOA), which provides a reference for the application of Tuina in KOA animal models. This protocol also studies the effective mechanism of Tuina for KOA, and the results will help promote its application.

Knee osteoarthritis (KOA), a common degenerative joint disorder, is characterized by chronic pain and disability, which can progress to irreparable ...

Tuina Manipulation to Reduce Inflammation and Cartilage Loss in Knee Osteoarthritis Rats

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