Clinically relevant aspects of stem cell technologies: current state of play

The emergence of stem cell technologies and their potential applications in regenerative medicine have generated immense interest by both the lay public and clinicians. Unproven and unregulated cell‐based therapies are commercially available both in Australia and internationally, and reports of patient uptake (stem cell tourism) and associated morbidity are increasingly frequent. Clinicians in all fields will require an enhanced understanding of the basic science principles and current state of play in regenerative medicine in order to effectively counsel patients regarding these therapies in the setting of both commercial ventures and clinical trials. This review aims to concisely highlight the key clinically pertinent features of these trials and briefly discuss the specific therapeutic potential, mechanism of action and risks associated with these variables.     

Nodular regenerative hyperplasia: a controversial indication for orthotopic liver transplantation

Nodular regenerative hyperplasia of the liver is an uncommon cause of portal hypertension. Patients with nodular regenerative hyperplasia have signs and symptoms of portal hypertension, without evidence of hepatocellular failure or encephalopathy. We report the case of a 44-year-old woman with recurrent esophageal bleeding and refractory ascites who had a history of hemosiderosis, hepatitis C, and chronic renal allograft rejection. Our preoperative diagnosis was cirrhotic end-stage liver disease and end-stage renal disease for which the patient underwent combined hepatic and renal transplantation. Her portal hypertension symptoms resolved, and her renal function has been normal for 18 months of follow-up. Histologic examination of the liver revealed nodular regenerative hyperplasia, and a review of the literature regarding the surgical management of patients with nodular regenerative hyperplasia revealed that various shunting procedures are generally recommended. After the failure of medical management in patients with nodular regenerative hyperplasia, portosystemic shunting may be indicated before proceeding to hepatic transplantation.     

Three Cases of Nodular Regenerative Hyperplasia of the Liver Following Renal Transplantation

We report three cases of nodular regenerative hyperplasia ofthe liver: the clinical onset of hepatic disease occurred between24 and 30 months after renal transplantation. Nodular regenerativehyperplasia was associated with peliosis hepatis in two cases,and with veno-occiusive disease in one case. Two patients developedportal hypertension, but are doing well. The third patient developedjaundice and died of septic shock. We discuss the aetiologicalrole of renal transplantation, cytomegalovirus infection, andazathioprine in the development of nodular regenerative hyperplasiaof the liver.     

Efficacy of a poly glycolic acid (PGA)/collagen composite nanofibre scaffold on cell migration and neovascularisation in vivo skin defect model

Abstract

Application of tissue engineering currently provides promising therapeutic options in the fields of plastic surgery and wound management. The ability of scaffold material for cell proliferation and differentiation is the key for tissue engineering. This study has developed a novel nanofibre composed of poly glycolic acid (PGA) and collagen, both of which have their own respective beneficial properties. This study aimed to estimate the in vivo efficiency of the PGA/collagen nanofibre on granulation histology and its ability to induce neovascularisation. The electrospinning technique produced the PGA/collagen nanofiber with a diameter of 500 nm and weight mixing ratio of 40%. The skin defects on the mouse model were covered with PGA/collagen or a commercially available collagen matrix (n = 9). The PGA/collagen group histologically showed significantly higher cell density and a fine microstructure with greater number of migrating cells as compared to collagen matrix. Then, both materials were applied to the microcirculatory angiogenesis model. The PGA/collagen group (n = 8) revealed significantly higher functional capillary density on days 5 and 7 after application. The findings substantiated the fact that our material had a superior ability regarding cellular migration and induction of neovascularisation compared with the elementary collagen matrix product. This better result might be attributed to the nano-size effect of fine structure and the incorporation of PGA, which has been associated with enhanced angiogenesis.     

Plasticity of spermatogonial stem cells

There have been significant breakthroughs over the past decade in the development and use of pluripotent stem cells as a potential source of cells for applications in regenerative medicine. It is likely that this methodology will begin to play an important role in human clinical medicine in the years to come. This review describes the plasticity of one type of pluripotent cell, spermatogonial stem cells (SSCs), and their potential therapeutic applications in regenerative medicine and male infertility. Normally, SSCs give rise to sperm when in the testis. However, both human and murine SSCs can give rise to cells with embryonic stem (ES) cell-like characteristics that can be directed to differentiate into tissues of all three embryonic germ layers when placed in an appropriate inductive microenvironment, which is in contrast to other postnatal stem cells. Previous studies have reported that SSCs expressed an intermediate pluripotent phenotype before differentiating into a specific cell type and that extended culture was necessary for this to occur. However, recent studies from our group using a tissue recombination model demonstrated that SSCs differentiated rapidly into another tissue, in this case, prostatic epithelium, without expression of pluripotent ES cell markers before differentiation. These results suggest that SSCs are capable of directly differentiating into other cell types without going through an intermediate ES cell-like stage. Because SSCs do not require reprogramming to achieve a pluripotent state, they are an attractive source of pluripotent cells for use in regenerative medicine.     

Is tissue engineering and biomaterials the future for lower urinary tract dysfunction (LUTD)/pelvic organ prolapse (POP)?

The fields of tissue engineering and regenerative medicine have seen major advances over the span of the past two decades, with biomaterials playing a central role. Although the term “regenerative medicine” has been applied to encompass most fields of medicine, in fact urology has been one of the most progressive. Many urological applications have been investigated over the past decades, with the culmination of these technologies in the introduction of the first laboratory‐produced organ to be placed in a human body. 1 With the quality of life issues associated with urinary incontinence, there is a strong driver to identify and introduce new technologies and the potential exists for further major advancements from regenerative medicine approaches using biomaterials, cells or a combination of both. A central question is why use biomaterials? The answer rests on the need to make up for inadequate or lack of autologous tissue, to decrease morbidity and to improve long‐term efficacy. Thus, the ideal biomaterial needs to meet the following criteria: (1) Provide mechanical and structural support, (2) Maintain compliance and be biocompatible with surrounding tissues, and (3) Be “fit for purpose” by meeting specific application needs ranging from static support to bioactive cell signaling. In essence, this represents a wide range of biomaterials with a spectrum of potential applications, from use as a supportive or bulking implant alone, to implanted biomaterials that promote integration and eventual replacement by infiltrating host cells, or scaffolds pre‐seeded with cells prior to implant. In this review we shall discuss the structural versus the integrative uses of biomaterials by referring to two key areas in urology of (1) pelvic organ support for prolapse and stress urinary incontinence, and (2) bladder replacement/augmentation. Neurourol. Urodynam. Neurourol. Urodynam. 30:775–782, 2011. © 2011 Wiley‐Liss, Inc.     

Low-intensity shock wave therapy for the treatment of vasculogenic erectile dysfunction: a narrative review of technical considerations and treatment outcomes

Erectile dysfunction (ED) impacts a significant portion of the aging male population. Standard treatments such as oral medications, intracavernosal injections, intraurethral suppositories, vacuum erection aids, and penile prosthesis placement have stood the test of time. Recently, there has been a growing interest in the concept of regenerative medicine with the goal of restoring or renewing functional tissue. Low intensity shock wave therapy (LiSWT) is one example of a regenerative therapy. A strong body of basic science data suggests that shockwaves, when applied to local tissue, will encourage blood vessel and nerve regeneration. Clinical evidence supports the use of LiSWT to treat conditions ranging from ischemic heart disease, musculoskeletal injuries, and even chronic non-healing wounds. LiSWT is also being used to treat male sexual dysfunction conditions such as Peyronie’s Disease and ED. The first studied application of LiSWT for ED was published in 2010. Since then multiple randomized, sham-controlled trials have sought to evaluate outcomes for this novel therapy in men with vasculogenic ED. Additionally, several meta-analyses are available with pooled data suggesting that LiSWT results in a significantly greater improvement in erectile function relative to sham-control. Despite these promising findings, the current body of literature is marred by significant heterogeneity relating to treatment protocols, patient populations, and follow-up duration. Further work is necessary to determine optimal device technologies, patient characteristics, and treatment duration prior to considering LiSWT as standard of care for men with ED.     

肝硬化非典型再生性结节的诊断及治疗

目的探讨肝硬化非典型再生性结节（ARN）的诊断及治疗。方法对24例经组织病理学证实ARN的病例资料进行回顾性的分析。结果本组病人，男15例，女9例，所有病人血清HbsAg（＋）；2例（8．69％）血清AFP（＋）。单个结节22／24（91．65％），两个结节1／24（4．17％）（均位于一侧肝叶内）。超声下边界清及有假包膜的18／24（75．0％）；多普勒下见动脉血供者1／24（4．17％）；CT：由于ARN以门静脉供血门静脉期增强者是20／24（83．33％）；MRI：T1WI上高信号15／24（62．5％）；T2WI上高信号3／24（12．5％）；术前超声定位下穿刺活检提示ARN者12／24（50％）。24例病人中：7例经B超引导下射频消融治疗；16例经手术切除，术后全部病人无严重并发症及死亡。结论ARN虽然属于良性病变，但是晚期肝硬化向肝细胞癌发展的一种重要组织学改变，临床上缺乏典型症状，多在影像学检查中发现，超声造影可以取代CT、MRI成为一种有效的检查手段。对于诊断明确的DN建议射频消融治疗并长期随访观察，但对于不能除外恶变者应该尽早手术切除。     

Diagnosing regenerative nodular hyperplasia, the “great masquerader” of liver tumors

Distinguishing benign tumors and pseudotumors of the liver from malignant tumors is a common clinical problem. Regenerative nodular hyperplasia (RNH) represents one of the more challenging pseudotumors to diagnose, because they can appear clinically indistinguishable from either a primary or a secondary liver malignancy. Even after comprehensive radiologic evaluation and image-guided percutaneous biopsy, the diagnosis of RNH can remain elusive. We reviewed the pathophysiology of RNH and present five cases illustrating the limitations of percutaneous biopsy and the utility of laparoscopic wedge biopsy in establishing the diagnosis. All patients underwent a complete workup that included percutaneous biopsy. Patients with a nondiagnostic percutaneous biopsy underwent a laparoscopic wedge biopsy or anatomical resection. H&E, vimentin, trichrome, and reticulin staining as well as CD34 immunostaining were performed. Five patients were diagnosed with RNH between May 2002 and April 2004. Three had focal nodular disease, whereas the other two had a diffuse multinodular presentation. Percutaneous biopsy definitively made the diagnosis in only one out of the five cases. Laparoscopic wedge biopsy was necessary to accurately make the diagnosis in three cases, whereas the fifth diagnosis was established after an anatomical resection. RNH is a unique pseudotumor of the liver that can present either as a solitary nodule or as a multinodular process. Percutaneous biopsy is associated with limitations in diagnosing RNH, and a more definitive surgical biopsy may be required. When RNH is considered, laparoscopic wedge biopsy is a safe and efficient way to obtain enough tissue to preserve the hepatic architecture required for diagnosis, while avoiding the morbidity of an unnecessary open resection.     

In vitro differentiation of progenitor cells isolated from juvenile pig hearts – expression of relevant gene and protein markers

Objectives. Heart failure is a significant cause of mortality worldwide, and most current therapies treat only its symptoms. The results of cardiac stem cell research suggest a promising treatment option for heart failure, but there is currently an unmet demand for better research models. We have therefore, for the first time, isolated, expanded and differentiated progenitor cells obtained from juvenile pig hearts to use as a platform for cardiac stem cell research. Design. Progenitor cells were isolated from the left ventricles of porcine hearts using collagenase enzymatic digestion and Percoll^®-gradient centrifugation. Cells were proliferated in Matrigel^®-coated wells. Cell differentiation was initiated by applying 5-azacytidine and subsequently controlled by modifying the serum concentration. Western blotting and qPCR were used to determine protein and gene expression, respectively. Results. Cardiac-specific genes, from the following proteins: troponin I-3, and myosin-heavy-chain 7 were stably expressed during proliferation and differentiation. Connexin-43 was upregulated and Actinin alfa 2 was downregulated during differentiation. The immature-cardiomyocyte marker GATA binding protein 4 was stably expressed but with a decrease in expression at day 4 of differentiation. Smooth muscle actin decreased in expression and Von Willebrand factor were stably expressed during differentiation. Smooth muscle protein expression was documented but no expression of cardiac-specific proteins after differentiation was found. Conclusion. The isolated progenitor cells had key cardiac-lineage gene expression characteristics but they did not express cardiac-specific proteins. Smooth muscle protein was expressed confirming commitment to the smooth muscle lineage.     

Quantitative analysis of intervertebral disc degeneration using Q-space imaging in a rat model

The degree of intervertebral disc (IVD) degeneration is qualitatively evaluated on T2-weighted imaging (T2WI). However, it is difficult to assess subtle changes in IVD degeneration using T2WI. Q-space imaging (QSI) is a quantitative diffusion-weighted imaging modality used to detect subtle changes in microenvironments. This study aimed to evaluate whether QSI can detect the inhibitory effects of the antioxidant N-acetylcysteine (NAC) in IVD degeneration. We classified female Wistar rats into control, puncture, and NAC groups (n = 5 per group). In the puncture and NAC groups, IVDs were punctured using a needle. The antioxidant NAC, which suppresses the progression of IVD degeneration, was orally administered in the NAC group 1 week prior to puncture. The progression and inhibitory effect of NAC in IVD degeneration were assessed using magnetic resonance imaging (MRI): IVD height, T2 mapping, apparent diffusion coefficient (ADC), and QSI. MRI was performed using a 7-Tesla system with a conventional probe (20 IVDs in each group). QSI parameters that were assessed included Kurtosis, the probability at zero displacement (ZDP), and full width at half maximum (FWHM). IVD degeneration by puncture was confirmed by histology, IVD height, T2 mapping, ADC, and all QSI parameters (P < .001); however, the inhibitory effect of NAC was confirmed only by QSI parameters (Kurtosis and ZDP: both P < .001; FWHM: P < .01). Kurtosis had the largest effect size (Kurtosis: 1.13, ZDP: 1.06, and FWHM: 1.02) when puncture and NAC groups were compared. QSI has a higher sensitivity than conventional quantitative methods for detecting the progressive change and inhibitory effect of NAC in IVD degeneration.     

Is Nodular Regenerative Hyperplasia of the Liver Associated with Azathioprine Therapy After Renal Transplantation?

A 33-year-old man presented with hepatosplenomegaly and ascites26 months after receiving a successful renal transplant. Immunosuppressionconsisted of low-dose prednisolone and azathioprine. Biopsyshowed characteristic features of nodular regenerative hyperplasiaof liver (NRHL), a condition which may be related to azathioprinetherapy but has rarely been reported in association with renaltransplantation.     

From intricate to integrated: Biofabrication of articulating joints

Articulating joints owe their function to the specialized architecture and the complex interplay between multiple tissues including cartilage, bone and synovium. Especially the cartilage component has limited self‐healing capacity and damage often leads to the onset of osteoarthritis, eventually resulting in failure of the joint as an organ. Although in its infancy, biofabrication has emerged as a promising technology to reproduce the intricate organization of the joint, thus enabling the introduction of novel surgical treatments, regenerative therapies, and new sets of tools to enhance our understanding of joint physiology and pathology. Herein, we address the current challenges to recapitulate the complexity of articulating joints and how biofabrication could overcome them. The combination of multiple materials, biological cues and cells in a layer‐by‐layer fashion, can assist in reproducing both the zonal organization of cartilage and the gradual transition from resilient cartilage toward the subchondral bone in biofabricated osteochondral grafts. In this way, optimal integration of engineered constructs with the natural surrounding tissues can be obtained. Mechanical characteristics, including the smoothness and low friction that are hallmarks of the articular surface, can be tuned with multi‐head or hybrid printers by controlling the spatial patterning of printed structures. Moreover, biofabrication can use digital medical images as blueprints for printing patient‐specific implants. Finally, the current rapid advances in biofabrication hold significant potential for developing joint‐on‐a‐chip models for personalized medicine and drug testing or even for the creation of implants that may be used to treat larger parts of the articulating joint. © 2017 The Authors. Journal of Orthopaedic Research Published by Wiley Periodicals, Inc. on behalf of the Orthopaedic Research Society. J Orthop Res 35:2089–2097, 2017.

     

The fate of recipient-derived hepatocytes in sex-mismatched liver allograft following liver transplantation

BACKGROUND: 'Bone marrow-derived stem cells' have attracted great attention as potential candidates for liver-directed gene therapy and as a tool for regenerative medicine. However, the fate of these cells is not well-known. The aim of this present study was to investigate the fate of 'recipient-derived repopulated hepatocytes' in sex-mismatched liver allografts in individuals following liver transplantation during systematic longitudinally performed liver biopsies. METHODS: Paraffin-embedded sex-mismatched liver biopsy samples of nine recipients (male/female ratio 5/4; mean age: 39.7 yr) were reviewed. Double labeling with immunohistochemistry for hepatocytes and recipient-specific bone marrow-derived cells and fluorescence in-situ hybridization for visualizing X and Y chromosomes were performed. These slides were examined systematically using an image analyzer system (Olympus microscope; Cyto-Vision, Applied Imaging, Biosciences Centre, Newcastle, UK). Only cells with two nuclear spots were considered for interpretation. RESULTS: The mean times from transplantation to first biopsy and between the first and the second biopsies were 5.9 and 20.9 months respectively. The proportion of recipient-derived repopulated hepatocytes was significantly decreased in the late biopsies when compared with the early biopsies (p = 0.001). All nine samples of the first biopsies had demonstrated recipient-derived hepatocyte repopulation, with a mean of 2.0%, whereas only seven of nine samples of the second biopsies had demonstrated recipient-derived hepatocyte repopulation with a low mean of 0.5% (p = 0.001). CONCLUSION: Based on these results, we suggest that 'recipient-specific bone marrow-derived hepatocyte repopulation' in liver allograft during tissue injury is a relatively early event.     

Rotator cuff tear state modulates self‐renewal and differentiation capacity of human skeletal muscle progenitor cells

Full thickness rotator cuff tendon (RCT) tears have long‐term effects on RC muscle atrophy and fatty infiltration, with lasting damage even after surgical tendon repair. Skeletal muscle progenitor cells (SMPs) are critical for muscle repair in response to injury, but the inability of RC muscles to recover from chronic RCT tear indicates possible deficits in repair mechanisms. Here we investigated if muscle injury state was a crucial factor during human SMP expansion and differentiation ex vivo. SMPs were isolated from muscles in patients with no, partial‐thickness (PT), or full‐thickness (FT) RCT tears. Despite using growth factors, physiological niche stiffness, and muscle‐mimetic extracellular matrix (ECM) proteins, we found that SMPs isolated from human RC muscle with RCT tears proliferated slower but fused into myosin heavy chain (MHC)‐positive myotubes at higher rates than SMPs from untorn RCTs. Proteomic analysis of RC muscle tissue revealed shifts in muscle composition with pathology, as muscle from massive RCT tears had increased ECM deposition compared with no tear RC muscle. Together these data imply that the remodeled niche in a torn RCT primes SMPs not for expansion but for differentiation, thus limiting longer‐term self‐renewal necessary for regeneration after surgical repair. © 2016 Orthopaedic Research Society. Published by Wiley Periodicals, Inc. J Orthop Res 35:1816–1823, 2017.

     

Autologous fat grafting efficacy in treating PostMastectomy pain syndrome: A prospective multicenter trial of two Senonetwork Italia breast centers

Postmastectomy pain syndrome (PMPS) represents a common complication following breast surgery defined as a chronic neuropathic pain located in the front of the chest, in the axilla and in the upper arm that for more than 3 months after surgery. Several medications prove to be ineffective while autologous fat grafting revealed to be an innovative solution in the treatment of neuropathic pain syndromes based on retrospective studies. For this reason, we performed a prospective multicenter trial to reduce the memory bias and further increase the evidence of the results. From February 2018 to March 2019, 37 female patients aged between 18 and 80 years, underwent mastectomy or quadrantectomy with pathologic scarring and chronic persistent neuropathic pain, compatible with PMPS, are been included in the study and treated with autologous fat grafting. During the enrollment phase, patients were asked to estimate pain using the Visual Analogue Scale (VAS) and POSAS questionnaire in order to evaluate scar outcomes. The VAS scale, starting from 6.9 (1.3), decreased in the first month by 3.10 (1.59), continuing to fall by 0.83 (1.60) to 3 months and by 0.39 (2.09) at 6 months. Statistical analysis showed a significant reduction after 1 month (P < .0001) and 3 months (P < .005). All POSAS grades documented a statistically significant reduction (P < .0001) of the scores by both observers and patients. We observed that no significant association was found between age, BMI, menopausal status of patients, days from oncologic surgery to autologous fat grafting and reduction of VAS values over time while both smoking and axillary dissection were observed as the main factor significantly associated with a reduced clinical efficacy (respectively, P = .0227 and P = .0066). Our prospective multicenter trial confirms the efficacy of fat grafting in the treatment of PMPS based on the principle of regenerative medicine with a satisfactory response in terms of pain reduction and improvement of the quality of the treated tissues. Clinical questionnaires show that the cicatricial areas improve in terms of color, thickness, skin pliability, and surface irregularities. Regenerative effect is based also on the adoption of needles. The combined effect of fat grafting and needles determines a clinical full response.