Protein-protein interactions (PPIs) are known to be involved in most cellular functions, and a detailed knowledge of such interactions is essential for studying their role in normal and pathological conditions. Significant progress is being made in the identification of PPIs through advances in computational methods. In particular, the AlphaFold2 machine learning-based model has been shown to accelerate drug discovery process by predicting the 3D structure of protein complexes. In this chapter, a straightforward protocol for predicting interprotein interactions between PAR-3 and its protein partner adapter molecule crk is provided. Such artificial intelligence-based and publicly available approaches can provide a resource for further investigation of therapeutic drug targets.

UNASSIGNED: Wilson\'s disease (WD) is not an uncommon genetic disease in clinical practice. However, the current WD therapies have limitations. The effectiveness of stem cell therapy in treating WD has yet to be verified, although a few animal studies have shown that stem cell transplantation could partially correct the abnormal metabolic phenotype of WD. In this case report, we present the therapeutic effect of human amniotic fluid containing stem cells in one WD patient.
UNASSIGNED: A 22-year-old Chinese woman was diagnosed with WD 1 year ago in 2019. The available drugs were not effective in managing the progressive neuropsychiatric symptoms. We treated the patient with pre-cultured human amniotic fluid containing stem cells. Amniotic fluid was collected from pregnant women who underwent induced labor at a gestational age of 19-26 weeks, and then, the fluid was cultured for 2 h to allow stem cell expansion. Cultured amniotic fluid that contained amniotic fluid derived stem cells (AFSC) in the range of approximately 2.8-5.5 × 104/ml was administrated by IV infusion at a rate of 50-70 drops per minute after filtration with a 300-mu nylon mesh. Before the infusion of amniotic fluid, low-molecular-weight heparin and dexamethasone were successively administrated. The patient received a total of 12 applications of amniotic fluid from different pregnant women, and the treatment interval depended on the availability of amniotic fluid. The neuropsychiatric symptoms gradually improved after the stem cell treatment. Dystonia, which included tremor, chorea, dysphagia, dysarthria, and drooling, almost disappeared after 1.5 years of follow-up. The Unified Wilson\'s Disease Rating Scale score of the patient decreased from 72 to 10. Brain magnetic resonance imaging (MRI) showed a reduction in the lesion area and alleviation of damage in the central nervous system, along with a partial recovery of the lesion to the normal condition. The serum ceruloplasmin level was elevated from undetectable to 30.8 mg/L, and the 24-h urinary copper excretion decreased from 171 to 37 μg. In addition, amniotic fluid transplantation also alleviates hematopoietic disorders. There were no adverse reactions during or after amniotic fluid administration.
UNASSIGNED: Amniotic fluid administration, through which stem cells were infused, significantly improves the clinical outcomes in the WD patient, and the finding may provide a novel approach for managing WD effectively.

Cell-to-cell communication must occur through molecular transport in the intercellular fluid space. Nanoparticles, such as exosomes, diffuse or move more slowly in fluids than small molecules. To find a microfluidic technology for real-time exosome experiments on intercellular communication between living cells, we use the microfluidic culture dish\'s quaternary ultra-slow microcirculation flow field to accumulate nanoparticles in a specific area. Taking stem cell-tumor cell interaction as an example, the ultra-slow microcirculatory flow field controls stem cell exosomes to interfere with tumor cells remotely. Under static coculture conditions (without microfluidics), the tumor cells near stem cells (<200 µm) show quick breaking through from its Matrigel drop to meet stem cells, but this \'breaking through\' quickly disappears with increasing distance. In programmed ultra-slow microcirculation, stem cells induce tumor cells 5000 μm far at the site of exosome deposition (according to nanoparticle simulations). After 14 days of programmed coculture, the glomeration and migration of tumor cells were observed in the exosome deposition area. This example shows that the ultra-slow microcirculation of the microfluidic culture dish has good prospects in quantitative experiments to study exosome communication between living cells and drug development of cancer metastasis.

The first product in the world for ex vivo cultivated oral mucosal epithelial cell transplantation (COMET) to treat limbal stem cell deficiency (LSCD), named Ocural®, was launched in June 2021 in Japan. COMET was performed on two patients, including the first case in the post-marketing phase of Ocural®. Pathological and immunohistochemical examinations were also carried out using specimens obtained before and after COMET and the spare cell sheet. In case 1, the ocular surface remained free from epithelial defects for approximately six months. In case 2, although defect of the cornea-like epithelia was observed after COMET for one month, it was resolved after the insertion of lacrimal punctal plugs. In case 1, adjuvant treatment was interrupted due to an accident during the second month after COMET, resulting in conjunctival ingrowth and corneal opacity. Eventually, a lamellar keratoplasty was required at six months after COMET. Immunohistochemistry revealed the presence of markers for stem cells (p63, p75), proliferation (Ki-67), and differentiation (Keratin-3, -4, and -13) in both the cornea-like tissue after COMET and a cultivated oral mucosal epithelial cell sheet. In conclusion, Ocural® can be accomplished without major complications, and the stem cells derived from oral mucosa might be successfully engrafted.

BACKGROUND: Asthma is a heterogeneous disorder characterized by chronic airway inflammation resulting in obstructive pulmonary symptoms. In preclinical studies, mesenchymal stem cells (MSCs) have demonstrated the ability to ameliorate the symptoms and immunologic pathways seen in asthma. Due to the known relationship between asthma and the hyper-responsive immune cascade, we hypothesized that MSCs could be an effective treatment option for patients with asthma due to their significant immunomodulatory properties.
OBJECTIVE: We present the initial results for the first patient enrolled in a phase 1 clinical trial (Safety of Cultured Allogeneic Adult Umbilical Cord Derived Mesenchymal Stem Cell Intravenous Infusion for the Treatment of Pulmonary Diseases).
METHODS: A 68-year-old male with a longstanding history of asthma requested mesenchymal stem cell treatment for his persistent asthma symptoms. Cultured umbilical cord-derived mesenchymal stem cells were infused intravenously at a dose of 100 million cells over a period of 40 minutes. Post-treatment follow- up was performed after two and six months.
RESULTS: The patient had no adverse events or complications related to treatment. In the two months posttreatment, his usage of a rescue inhaler decreased to 1 time per month, over 90% reduction. In addition, he had a 70% reduction in nebulizer usage. Improvement was sustained in the 6 months follow-up.
CONCLUSIONS: We report the first case of mesenchymal stem cell treatment significantly and safely improving asthma clinical symptoms in a human. Additionally, an extensive literature review provided several plausible mechanisms by which stem cells can ameliorate immune hyper-stimulation associated with asthma.

BACKGROUND: Back pain causes tremendous patient suffering and high financial cost to the healthcare system. Mesenchymal Stem Cells (MSCs) have demonstrated the ability to enhance healing and reduce inflammation and pain without the deleterious side effects of corticosteroids and nonsteroidal anti-inflammatory drugs in numerous clinical series for peripheral joint arthritis. We hypothesized that translaminar MSC injection into the epidural space would effectively treat disc arthritis without the burden of sedation and the risks of disc space injection. We further hypothesized that MSC injection into the facet joints would effectively and safely treat facet joint-induced back pain. The combination of epidural and facet joint injection would potentially treat the most recognized low back pain generators with virtually complete safety.
OBJECTIVE: We present the initial results for the first patient enrolled in phase 1 clinical trial of the efficacy and safety of allogeneic MSCs when injected translaminarly and into the facet joints for the treatment of recalcitrant discogenic and arthritic back pain.
METHODS: A 47-year-old male presented with complaints of 13-year-long chronic lower back pain resistant to conservative treatment. The decision was made to treat the patient with umbilical cord-derived MSCs. 87 million MSCs were infused intravenously. Simultaneously, 1 million cells were injected into each of the 8 lumbar epidural facet joints and 5 million cells into the lumbar epidural space. The patient had no adverse events or complications related to the treatment. Five days after treatment, most of his lumbar pain was gone, and his back spasms stopped. He no longer needed to take acetaminophen or ibuprofen and had no difficulty sleeping without medications. The patient also reported his residual cervical radicular pain to be 98% resolved due to the injection.
CONCLUSIONS: We have demonstrated for the first time that MSC injection into the lumbar facet joints and epidural space results in significant improvement of lower back pain and can improve symptoms in other spinal regions without engendering the risks associated with intradiscal injections or epidural use of corticosteroids.

Acute lymphoblastic leukemia (ALL) is the most frequent childhood cancer, with 80-85% represented by B cell ALL and only 15% by T cell ALL. T Cell ALL (T-ALL) carries a more reserved prognosis compared to B Cell ALL (B-ALL) with regard to response to treatment, risk of relapse, and overall survival. Progress made in current monitoring protocols such as via flow cytometry immunophenotyping (FCM) and by PCR-based amplification of antigen-receptor genes led to improved management of patients with ALL and superior rates of survival. Nevertheless, challenges remain in some clinical cases. This manuscript describes a unique case of T-ALL and raises awareness of such clinical challenges. The article presents an overview of the flow cytometry immunophenotyping at diagnosis and during treatment of a pediatric patient with T-ALL from Fundeni Clinical Institute. In this case, in spite of various therapeutic measures such as first-line chemotherapy for high risk group, salvage chemotherapy (FLAG), conditioning regimen (FLU-BU-TT-ATG), and stem cell transplant, a chemoresistance clone continued to be present.

En coup de sabre deformity (ECDS) is a form of localized scleroderma in the frontoparietal region caused by progressive subcutaneous tissue atrophy and bony defect. Although ECDS involves two layers, skin/subcutaneous tissue and bone, the existing literature mainly focuses only on treating the skin/subcutaneous tissue layer. In this case series, we aimed to propose a novel approach that includes the combined use of fat grafting and demineralized bone matrix (DBM). Four patients with ECDS deformity, operated between February 2016 and October 2018, were retrospectively evaluated. All the patients were treated with the novel approach. Patients were evaluated with localized scleroderma scale and computed tomography (CT) scan in the preoperative period and at the annual follow-up. We observed remarkable improvement in the localized scleroderma scale including appearance, palpation, and size scores in all patients at the annual follow-up. CT scans at the annual follow-up revealed new callus formation at the bony defect area in all patients. Reinforcing fat grafting with DBM could promote healing of the bony and skin/subcutaneous tissue defects associated with ECDS.

BACKGROUND: With the advent of innovative therapies including biologics and Janus kinase inhibitors, children with rheumatic diseases are more likely to have improved outcomes. Despite these advances, some children do not respond, or they, or their parents fear adverse events and seek other alternatives. Increasingly, private companies are offering mesenchymal stem cells (MSC) as an alternative, which are described as natural therapies for rheumatic diseases, often insinuating them as a cure. MSC have immunomodulatory properties, and transplantation of these stem cells have been used to successfully treat immunologic conditions like graft-versus-host disease. Lately, MSC research in adult lupus has been encouraging, but the clinical trials are still underway and in most, MSC therapy is not a standalone treatment. This retrospective case series will highlight three cases of pediatric refractory autoimmune disease whose parents sought out and received MSC therapy as a self-decision without first seeking medical advice from our specialty. The three families felt that their children were improved and in two believed that their child was cured. MSC have the potential of beneficial immunomodulation and may be a powerful tool in the therapy of rheumatic disease, but well controlled clinical trials are necessary and should be designed and monitored by experts in childhood rheumatic disease.
METHODS: Three children with three different rheumatic diseases; systemic lupus erythematosus, mixed connective tissue disease and juvenile idiopathic arthritis were under the care of pediatric rheumatology at a large, tertiary-care, teaching institution. Multiple non-biologic and biologic disease-modifying anti-rheumatic drugs failed to significantly decrease disease activity, and as a result, the families chose to undergo MSC therapy. After transplantation, all children improved per patient and parent report and tapered off conventional immunosuppressive drugs. No serious adverse events occurred in these three patients.
CONCLUSIONS: The three cases presented in this report reflect comparable beneficial outcomes and minimal risks published in adult studies. These were not controlled studies, however, and benefit was reported rather than documented. These cases suggest that MSC transplantation may prove a promising adjunctive treatment option; however, further research, development of standardized infusion therapy protocols, and well-designed monitored clinical trials are essential.

Background. Large mandibular defects are considered difficult reconstructive challenges for oral and maxillofacial surgeons. Cell therapy, as an alternative technique, might increase the speed of bone regeneration. This study aimed to investigate bone regeneration in large defects of dog mandibles using allogenic adipose-derived stem cells on gelatin foam as a cell carrier. Methods. The tissue engineering phase consisted of the sampling of adult dogs\' adipose tissue that can easily be isolated from adipose stem cells (ASCs) of the dogs, ASCs were cultured in Dulbecco\'s Modified Eagle\'s Medium (DMEM, Gibco, USA) with low glucose, containing 10% fetal bovine serum (FBS) (Sigma, USA) and 1% penicillin-streptomycin (Gibco, USA), with the characterization of dog ASCs and gelatin-transplanted ASCs. Six dogs were included in this experimental study in the next step and randomly assigned to the treatment and control groups. The samples in both groups underwent surgery under general anesthesia to create uniform 3-cm bony defects. The samples in both groups were reconstructed with titanium reconstruction plates and screws. A large bone gap filled with ASCs (5×106 ) was seeded on gelatin (ASCs) in the treatment group. In the control group, bony defects were filled with a cell delivery carrier without ASCs. Six months after transplantation, the animals\' mandibles were evaluated by CT scan imaging, and the results were quantified through the Hounsfield unit (HU). The data were analyzed with t-test. Results. Before transplantation, the nature of the stem cells was confirmed by the expression of CD44 and CD105 cell markers at 71.9% and 89.3%, respectively, and a lack of the CD45 cell marker expression at 2.2%. Evaluation of CT scan images showed significantly higher bone repair in the ASCs group (920.25±572.92 HU) than in the control group (-94.746± 08.42). Conclusion. The bone regeneration of the ASCs group was significantly higher than that in the control group.