Development and validation of magnetic bead pentaplex immunoassay for simultaneous quantification of murine serum IgG antibodies to acellular pertussis, diphtheria and tetanus antigens used in combination vaccines.

We describe here a magnetic bead-based multiplex (pentaplex) immunoassay (MIA) platform developed as an alternative to enzyme-linked immunosorbent assays (ELISA) used in immunogenicity testing of DTaP/TdaP vaccine in animals. MIA simultaneously measures the concentration of serum (IgG) antibodies against B. Pertussis antigens; pertussis toxin, filamentous hemagglutinin (FHA), pertactin (PRN) and tetanus (T) and diphtheria (D) toxoid in the Tdap vaccine immunized animals. Assay validation experiments were done using a panel of serum samples. The results are expressed in IU/ml using WHO reference mice serum. The standard curve was linear with 4PL logistic fit over an eight 2-fold dilution range with LOQ of 0.003, 0.022, 0.005 IU/ml for PT, FHA and PRN and 0.016 U/ml for T and D antigens indicating sensitivity. No interference was observed in monoplex versus multiplex measurements. Specificity was demonstrated by ≥90% homologous and ≤15% heterologous inhibition for all the antigens. The assay was reproducible, with a mean coefficient of variation (CV) of ≤10% for intra-assay duplicates and ≤25% for interassays using different lots of beads and analyst. Accuracy was demonstrated wherein the ratio of observed vs. assigned unitages were within 80-120%. The study suggests that the Pentaplex (MIA) platform meets all the criteria for the serological assay combination vaccines with additional advantages of high throughput, reduced sample volumes, faster analysis with reduced manpower in contrast to conventional monoplex ELISA.

Mechanisms, Efficacy, and Clinical Applications of Platelet-Rich Plasma in Tendinopathy: A Comprehensive Review.

Tendinopathy poses a significant clinical challenge characterized by chronic tendon pain, swelling, and impaired function, affecting athletes and the general population. Current treatments often provide limited success, necessitating exploration into regenerative therapies such as platelet-rich plasma (PRP). PRP harnesses the regenerative potential of autologous platelets and growth factors to promote tendon healing. This review aims to comprehensively examine the mechanisms, efficacy, and clinical applications of PRP in tendinopathy. We discuss the pathophysiology of tendinopathy, highlighting collagen disorganization, increased ground substance, and inflammatory changes. PRP's action mechanism involves releasing bioactive molecules that stimulate cellular proliferation, collagen synthesis, and tissue remodeling. Clinical studies and trials evaluating PRP in various tendinopathies, including Achilles, patellar, and rotator cuff tendinopathy, are reviewed to assess its efficacy and effectiveness compared to traditional therapies. Practical aspects, such as preparation methods, injection techniques, and safety considerations, are discussed to provide insights into optimal PRP administration. Challenges, including protocol variability and evidence gaps, are addressed, and future research and clinical practice directions are proposed. By synthesizing current knowledge, this review aims to guide clinicians in enhancing treatment strategies and advancing the field of tendon regenerative medicine.

Therapeutic Potential of Platelet-Rich Plasma in Fracture Healing: A Comprehensive Review.

Fracture healing is a dynamic process essential for the restoration of bone integrity and function. However, factors such as patient age, comorbidities, and the severity of the fracture can impede this process, leading to delayed healing or nonunion. Platelet-rich plasma (PRP) has emerged as a promising therapeutic option for enhancing fracture healing. PRP is an autologous blood product containing a concentrated mixture of platelets, growth factors, and cytokines known to promote tissue regeneration and repair. This comprehensive review provides an overview of the fracture healing process, emphasizing the importance of timely and efficient bone repair. We discuss the mechanisms underlying the purported efficacy of PRP in fracture healing, drawing upon both preclinical and clinical evidence. Preclinical studies in animal models have demonstrated the ability of PRP to accelerate fracture healing, stimulate osteogenesis, and enhance bone regeneration. Clinical studies have yielded mixed results, with some reporting positive outcomes in terms of accelerated healing and improved functional outcomes, while others have shown no significant benefits over standard treatments. Factors influencing the efficacy of PRP, such as timing of administration, PRP concentration, and patient-specific variables, are also examined. Furthermore, safety considerations and potential adverse effects associated with PRP therapy are discussed. Despite the promising preclinical findings, challenges remain in standardizing PRP formulations, optimizing administration protocols, and addressing unanswered questions regarding its long-term efficacy and safety. This review aims to provide insights into the therapeutic potential of PRP in fracture healing, informing future research directions and guiding clinical practice.

Navigating the Therapeutic Landscape: A Comprehensive Review of Platelet-Rich Plasma and Bone Marrow Aspirate Concentrate in Knee Osteoarthritis.

This comprehensive review provides an in-depth analysis of platelet-rich plasma (PRP) and bone marrow aspirate concentrate (BMAC) as potential treatments for knee osteoarthritis. It explores their mechanisms of action, clinical efficacy, safety considerations, and the importance of personalised treatment approaches. The review highlights promising findings regarding the ability of PRP and BMAC to alleviate symptoms, improve joint function, and potentially slow disease progression. It emphasises the need for further research into long-term outcomes, direct comparative studies, protocol standardisation, biomarker identification, and cost-effectiveness assessments to enhance clinical practice. While the review does not directly compare PRP and BMAC, it provides valuable insights into their respective roles in knee osteoarthritis management. The review aims to contribute to evidence-based advancements in regenerative therapies for knee osteoarthritis by addressing critical research priorities and refining treatment strategies.

Harnessing Healing Power: A Comprehensive Review on Platelet-Rich Plasma in Compound Fracture Care.

This comprehensive review explores the applications of platelet-rich plasma (PRP) in the context of compound fracture care, providing a thorough examination of its biological mechanisms, preparation techniques, and clinical implications. The analysis highlights PRP's potential in accelerating bone healing, enhancing soft tissue repair, reducing inflammation and infection risks, and managing pain during fracture recovery. The review underscores the importance of ethical and regulatory considerations in integrating PRP into orthopaedic practice, emphasising informed consent, transparent patient communication, and ongoing monitoring of ethical concerns. Looking ahead, the implications for the future of compound fracture care suggest a transformative shift with the potential for personalised medicine approaches and emerging technologies. However, the conclusion calls for a balanced perspective, acknowledging the promising applications of PRP while emphasising the need for responsible and ethical use. The collaborative efforts of healthcare professionals, researchers, and regulatory bodies are crucial in navigating this evolving landscape and harnessing the healing power of PRP to redefine orthopaedic care for individuals with compound fractures.

Ayurveda-based Botanicals as Therapeutic Adjuvants in Paclitaxel-induced Myelosuppression.

Chemotherapy-induced myelosuppression is one of the major challenges in cancer treatment. Ayurveda-based immunomodulatory botanicals Asparagus racemosus Willd (AR/Shatavari) and Withania somnifera (L.). Dunal (WS/Ashwagandha) have potential role to manage myelosuppression. We have developed a method to study the effects of AR and WS as therapeutic adjuvants to counter paclitaxel (PTX)-induced myelosuppression. Sixty female BALB/c mice were divided into six groups-vehicle control (VC), PTX alone, PTX with aqueous and hydroalcoholic extracts of AR (ARA, ARH) and WS (WSA, WSH). The myelosuppression was induced in mice by intraperitoneal administration of PTX at 25 mg/kg dose for three consecutive days. The extracts were orally administered with a dose of 100 mg/kg for 15 days prior to the induction with PTX administration. The mice were observed daily for morbidity parameters and were bled from retro-orbital plexus after 2 days of PTX dosing. The morbidity parameters simulate clinical adverse effects of PTX that include activity (extreme tiredness due to fatigue), behavior (numbness and weakness due to peripheral neuropathy), body posture (pain in muscles and joints), fur aspect and huddling (hair loss). The collected samples were used for blood cell count analysis and cytokine profiling using Bio-Plex assay. The PTX alone group showed a reduction in total leukocyte and neutrophil counts (4,800 ± 606; 893 ± 82) when compared with a VC group (9,183 ± 1,043; 1,612 ± 100) respectively. Pre-administration of ARA, ARH, WSA, and WSH extracts normalized leukocyte counts (10,000 ± 707; 9,166 ± 1,076; 10,333 ± 1,189; 9,066 ± 697) and neutrophil counts (1,482 ± 61; 1,251 ± 71; 1,467 ± 121; 1,219 ± 134) respectively. Additionally, higher morbidity score in PTX group (7.4 ± 0.7) was significantly restricted by ARA (4.8 ± 1.1), ARH (5.1 ± 0.6), WSA (4.5 ± 0.7), and WSH (5 ± 0.8). (Data represented in mean ± SD). The extracts also significantly modulated 20 cytokines to evade PTX-induced leukopenia, neutropenia, and morbidity. The AR and WS extracts significantly prevented PTX-induced myelosuppression (p < 0.0001) and morbidity signs (p < 0.05) by modulating associated cytokines. The results indicate AR and WS as therapeutic adjuvants in cancer management.

Estrogen upregulates inflammatory signals through NF-κB, IFN-γ, and nitric oxide via Akt/mTOR pathway in the lymph node lymphocytes of middle-aged female rats.

The alterations in the secretion of sex steroids, especially estrogen, in females throughout reproductive life and its decline with age alters the functions of the neuroendocrine-immune network and renders them susceptible to age-related diseases and cancers. This study investigates the mechanisms of estrogen-induced alterations in cell-mediated immune and inflammatory responses in the lymphocytes from lymph nodes (axillary and inguinal) of ovariectomized (OVX) middle-aged female rats. Ovariectomized middle-aged (MA) Sprague-Dawley female rats (n=8) were implanted with 17ß-estradiol (E2) 30-day release pellets (0.6 and 300µg). At the end of the treatment period, lymph nodes (axillary and inguinal) were isolated and examined for serum 17ß-estradiol, lymphoproliferation, cytokine production, expression of p-Akt, p-mTOR, p-IκB-α and p-NF-κB (p50 and p65), extent of lipid peroxidation, nitric oxide (NO) production, cytochrome c oxidase activity and reactive oxygen species (ROS) production. There was an OVX-related decline in serum 17ß-estradiol level, Con A-induced lymphoproliferation, p-Akt and p-mTOR expression, and cytochrome c oxidase (COX) activity. E2 supplementation increased serum 17ß-estradiol level, lymphoproliferation, expression of p-Akt, p-mTOR, p-IκB-α and p-NF-κB (p50 and p65), lipid peroxidation, IFN-γ, TNF-α, ROS and NO production, while it decreased IL-6 production. E2 mediates inflammatory responses by increasing the levels of NO and TNF-α by up regulating IFN-γ and simultaneously promotes aging through the generation of free radicals as reflected by increased lipid peroxidation and ROS production in lymph nodes. These findings may have wide implications to immunity and inflammatory disorders including autoimmune diseases predominantly prevalent in females.

Estrogen modulates neural-immune interactions through intracellular signaling pathways and antioxidant enzyme activity in the spleen of middle-aged ovariectomized female rats.

Modulation of neural-immune interactions by estrogen in the spleens of ovariectomized (OVX) middle-aged female rats was examined. Con A-induced lymphoproliferation, splenic tyrosine hydroxylase (TH) and nerve growth factor (NGF) expression, levels of p-ERK 1/2, p-CREB, and p-Akt, and activity of superoxide dismutase decreased in OVX rats while estrogen treatment enhanced their expression, levels, and activity. Also, estrogen treatment enhanced Con A-induced IFN-γ production and decreased Con A-induced IL-2 production compared to OVX animals. In contrast, estrogen increased the extent of lipid peroxidation and protein carbonyl formation while OVX induced a decline in protein carbonyl formation. These results suggest that estrogen enhances neural-immune interactions while simultaneously affecting it through generation of free radicals as reflected by increased lipid peroxidation and protein carbonyl formation.