[Show abstract][Hide abstract] ABSTRACT: Inflammatory response is one of the key components of pain perception. Continuous infusion (CWI) of local anesthetics has been shown to be effective in controlling pain and reducing postoperative morphine consumption, but the effect of adding a potent anti-inflammatory drug (such as a steroid) has never been addressed. In our study, we want to investigate the effect of CWI with local anesthetic + methylprednisolone on acute and persistent pain, correlating clinical data with biomarkers of inflammation and genetic background.
After approval by their institutional review board, three hospitals will enroll 120 patients undergoing major abdominal surgery in a randomized, double-blind, phase III study. After a 24-h CWI of ropivacaine 0.2 % + methylprednisolone 1 mg/kg, patients will be randomly assigned to receive either ropivacaine + steroid or placebo for the next 24 h. Then, patient-controlled CWI with only ropivacaine 0.2 % or placebo (according to the group of randomization) is planned after 48 h up to 7 days (bolus 10 ml, lock-out 1 h, maximum dose of 40 ml in 4 h). Morphine equivalent consumption up to 7 days will be analyzed, together with any catheter- or drug-related side effect. Persistent post-surgical pain (PPSP) incidence will also be investigated. Our primary endpoint is analgesic consumption in the first 7 days after surgery; we will evaluate, as secondary endpoints, any catheter- or drug-related side effect, genotype/phenotype correlations between some polymorphisms and postoperative outcome in terms of morphine consumption, development of the inflammatory response, and incidence of PPSP. Finally, we will collect, in a subgroup of patients, wound exudate samples by micro-dialysis, blood samples, and urine samples up to 72 h to investigate local and systemic inflammation and oxidative stress.
This is a phase III trial to evaluate the safety and efficacy of wound infusion with steroid and local anesthetic. The study is aimed also to evaluate how long this infusion has to be maintained in order to maximize effectiveness. Our data are intended to quantify the amount of ropivacaine and methylprednisolone needed by patients undergoing major abdominal surgery, to be stored in a new nanotechnology device for sustained pain treatment after surgery. We also aim to clarify the roles of inflammatory response, oxidative stress, and genetic background on postoperative and persistent pain after major abdominal surgery.
The trial was registered on ClinicalTrials.gov ( NCT02002663 ) on 24 Oct. 2013.
[Show abstract][Hide abstract] ABSTRACT: Mesenchymal stemcells (MSCs) have been proposed as a potential therapeutic tool for Parkinson’s disease (PD) and systemic administration of these cells has been tested in preclinical and clinical studies. However, no information on survival and actual capacity of MSCs to reach the brain has been provided. In this study,weevaluatedhoming of intraarterially infused ratMSCs (rMSCs) in thebrain of rats bearing a 6-hydroxydopamine (6-OHDA)-induced lesion of the nigrostriatal tract, to establishwhether the toxininduced damage is sufficient to grant MSC passage across the blood-brain barrier (BBB) or if a transient BBBdisruptionisnecessary.The rMSCdistributioninperipheralorgans and theeffects of cell infusionon neurodegenerative process andmotor deficitswere also investigated. rMSCswere infused 14 days after 6-OHDA injection. A hyperosmolar solution of mannitol was used to transiently permeabilize the BBB. Behavioralimpairmentwas assessedbyadjusting steptestandresponsetoapomorphine.Animalswere sacrificed 7 and 28 days after cell infusion. Our work shows that appreciable delivery of rMSCs to the brainof6-OHDA-lesionedanimalscanbeobtainedonlyaftermannitol pretreatment.Anotablepercentage of infused cells accumulated in peripheral organs. Infusion of rMSCs did notmodify the progression of 6-OHDA-induced damage or themotor impairment at the stepping test, but induced progressive normalization of the pathological response (contralateral turning) to apomorphine administration. These findings suggest thatmany aspects should be further investigated before considering any translation of MSC systemic administration into the clinical setting for PD treatment.
[Show abstract][Hide abstract] ABSTRACT: Mesenchymal stromal cells (MSCs) expanded in vitro have been proposed as a potential therapy for congenital or acquired skin defects in pediatrics. The aim of this pre-clinical study was to investigate the effects of intradermal injections of MSC in experimental cutaneous wound repair comparing allogeneic and autologous adipose stem cells (ASCs) and autologous bone marrow-mesenchymal stromal cells (BM-MSCs).
Mesenchymal stromal cells were in vitro expanded from adipose and BM tissues of young female New Zealand rabbits. MSCs were characterized for plastic adhesion, surface markers, proliferation and differentiation capacity. When an adequate number of cells (ASCs 10 × 10(6) and BM-MSCs 3 × 10(6), because of their low rate of proliferation) was reached, two skin wounds were surgically induced in each animal. The first was topically treated with cell infusions, the second was used as a control. The intradermal inoculation included autologous or allogeneic ASCs or autologous BM-MSCs. For histological examination, animals were sacrificed and wounds were harvested after 11 and 21 days of treatment.
Rabbit ASCs were isolated and expanded in vitro with relative abundance, cells expressed typical surface markers (CD49e, CD90 and CD29). Topically, ASC inoculation provided more rapid wound healing than BM-MSCs and controls. Improved re-epithelization, reduced inflammatory infiltration and increased collagen deposition were observed in biopsies from wounds treated with ASCs, with the best result in the autologous setting. ASCs also improved restoration of skin architecture during wound healing.
The use of ASCs may offer a promising solution to treat extended wounds. Pre-clinical studies are however necessary to validate the best skin regeneration technique, which could be used in pediatric surgical translational research.
Full-text · Article · Jul 2015 · Journal of Translational Medicine
[Show abstract][Hide abstract] ABSTRACT: In the last two decades, animal models have become important tools in understanding and treating pain, and in predicting analgesic efficacy. Although rodent models retain a dominant role in the study of pain mechanisms, large animal models may predict human biology and pharmacology in certain pain conditions more accurately. Taking into consideration the anatomical and physiological characteristics common to man and pigs (median body size, digestive apparatus, number, size, distribution and communication of vessels in dermal skin, epidermal-dermal junctions, the immunoreactivity of peptide nerve fibers, distribution of nociceptive and non-nociceptive fiber classes, and changes in axonal excitability), swines seem to provide the most suitable animal model for pain assessment. Locomotor function, clinical signs, and measurements (respiratory rate, heart rate, blood pressure, temperature, electromyography), behavior (bright/quiet, alert, responsive, depressed, unresponsive), plasma concentration of substance P and cortisol, vocalization, lameness, and axon reflex vasodilatation by laser Doppler imaging have been used to assess pain, but none of these evaluations have proved entirely satisfactory. It is necessary to identify new methods for evaluating pain in large animals (particularly pigs), because of their similarities to humans. This could lead to improved assessment of pain and improved analgesic treatment for both humans and laboratory animals.
Full-text · Article · May 2014 · Journal of Pain Research
[Show abstract][Hide abstract] ABSTRACT: Introduction and Aims: Hypertensive nephrosclerosis is one of the most frequent causes of chronic kidney failure. Proteomic analysis represents
one possibility to improve the pathophysiologic knowledge and diagnostic precision of this disorder. In this study, we investigated
experimental nephrosclerosis in two-kidney, one-clip (2K1C) hypertensive rats.
Methods: The renal cortex proteome from juxtamedullary cortex (JMC) and outer cortex (OC) of 2K1C male Hannover Wistar rats (n=4)
was compared to sham-operated controls (n=6) using mass spectrometry based quantitative proteomics. We combined a high abundant
plasma protein depletion strategy with an extended liquid chromatographic gradient to improve peptide and protein identification.
Immunohistology was used for confirmation of the abundance of a selected protein.
Results: In total we identified 1,724 proteins, of which 1,434 could be quantified based on ≥ 2 unique peptides. Comparative proteomics
revealed 683 proteins, including PDGFR-β pathway, with different abundance between the non-clipped renal cortex of hypertensive
2K1C rats and of the corresponding kidney in normotensive controls (p<0.05, absolute fold change ≥1.5). A total of 12 proteins
were differentially regulated between JMC and OC of 2K1C animals. Among the most significantly altered proteins in the whole
cortex, we identified periostin, transgelin and creatine kinase B-type with known association to renal fibrosis. Their relative
abundance pattern separates 2K1C and controls based on a 3D scatter plot (Fig. 1A) and on different decision algorithms (Table
1). Also, the relative abundance of periostin alone indicated perfect classification, as assessed by a scatter plot (Fig.
1B). Enrichment of periostin was verified by immunohistology showing no periostin staining in control cortex (Fig. 2A) but
clear positivity around fibrotic vessels in nephrosclerosis (Fig. 2B).
Conclusions: The proteome is substantially altered in hypertension-induced kidney damage. We propose periostin and especially periostin
in combination with transgelin and creatine kinase B-type as a possible proteomic classifier to distinguish hypertensive nephrosclerosis
from normal tissue. This classifier needs to be further validated with respect of early fibrosis diagnosis, of prognosis and
for its potential as a novel target.
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Full-text · Article · May 2014 · Nephrology Dialysis Transplantation
[Show abstract][Hide abstract] ABSTRACT: Introduction and Aims: IgA nephropathy, the most common primary glomerulonephritis worldwide, can lead to end-stage renal disease and kidney transplantation. Disease recurrence frequently occurs after transplantation. We investigated the predictive value of three markers including galactose-deficient (Gd) IgA1, IgG anti-IgA autoantibodies, IgA-soluble (s) CD89 complexes for IgA nephropathy recurrence. The efficacy of intravenous pulse steroid administration for treatment of recurrent IgA nephropathy was evaluated.Methods: The IgA nephropathy recurrent group (R group, n=11) was compared to matched patients transplanted for IgA nephropathy but without recurrence (NR group, n=13) and healthy subjects (n=22) for proportions of serum Gd-IgA1, IgA-IgG complexes and IgA-sCD89 complexes. Efficacy of pulse steroid therapy in reducing proteinuria was analysed in kidney transplant recipients R group.Results: Pre-transplantation serum proportion of Gd-IgA1 and IgA-IgG complexes were higher in R group compared to NR g
Full-text · Article · May 2014 · Nephrology Dialysis Transplantation
[Show abstract][Hide abstract] ABSTRACT: Introduction and Aims: Peritubular capillaritis (ptc), an engorgement of immune cells in the peritubular capillaries, is one of the diagnostic criteria
of antibody-mediated rejection (AMR). According to the Banff classification the severity of ptc, its distribution and its
cellular composition should be routinely reported. It remains unclear whether or to which extent the morphological presentation
of ptc is associated with clinical outcomes.
Methods: This retrospective study included 749 biopsied renal transplant recipients (transplantation 1999 - 2006; 1322 indication
biopsies). Detailed characterization of ptc included (i) its composition [mainly monocytic (>75% monocytes), granulocytic
(>75% granulocytes) or mixed], (ii) its distribution [focal (10-50% of renal cortex peritubular capillaries) vs. diffuse (>50
%)] and (iii) its severity scored according to the Banff scheme (ptc: 1,2,3), respectively. Biopsy results were analyzed in
relation to clinical outcomes.
Results: Ptc was present in 24.9% (n=329) of the studied allograft biopsies. Ptc scores 1, 2 and 3 were observed in 43%, 47% and 10%
of these specimens. Leucocyte subpopulations were categorized focal vs. diffuse monocytic (11%/24%), focal vs. diffuse granulocytic
(4%/4%), or focal vs. diffuse mixed (14%/20%). In 23% of the cases ptc was not classified (e.g. vicinity of tubular necrosis,
pyelonephritis or infarct). Ptc scores (1, 2 or 3) were associated with both diagnosis of AMR and cellular rejection (ACR).
Diffuse monocytic or mixed ptc (OR=4.51, 95% CI 1.82-11.16, and OR=6.8, 2.44-19.36, p≤0.001), but not granulocytic ptc (OR=3.81,
95% CI 0.41-35.02, p=0.2) were associated with AMR. At the same time, diffuse ptc was associated with ACR, regardless of its
composition (monocytic: OR=6.1, 95%CI 3.61-12.44; granulocytic: OR=6.69, 1.58-28.41; mixed: OR=8.05, 4.06-15.96; p≤0.01).
In uni- and multivariate analysis ptc 3 (OR=2.56, 95%CI 1.24-1.96, p=0.011), diffuse monocytic or mixed ptc (OR=1.75, 95%CI
1.01-3.06, p=0.048 and OR=1.75, 1.03-3.26, p=0.04) were risk factors for inferior graft survival.
Conclusions: Our results underscore a high relevance of the detailed qualitative and quantitative characterization of peritubular capillaritis
in indication biopsies as a predictor for inferior clinical outcomes.
Full-text · Article · May 2014 · Nephrology Dialysis Transplantation
[Show abstract][Hide abstract] ABSTRACT: In osteosarcoma (OS) and most Ewing sarcoma (EWS) patients, the primary tumor originates in the bone. Although tumor resection surgery is commonly used to treat these diseases, it frequently leaves massive bone defects that are particularly difficult to be treated. Due to the therapeutic potential of mesenchymal stem cells (MSCs), OS and EWS patients could benefit from an autologous MSCs-based bone reconstruction. However, safety concerns regarding the in vitro expansion of bone marrow-derived MSCs have been raised. To investigate the possible oncogenic potential of MSCs from OS or EWS patients (MSC-SAR) after expansion, this study focused on a biosafety assessment of MSC-SAR obtained after short- and long-term cultivation compared with MSCs from healthy donors (MSC-CTRL).
We initially characterized the morphology, immunophenotype, and differentiation multipotency of isolated MSC-SAR. MSC-SAR and MSC-CTRL were subsequently expanded under identical culture conditions. Cells at the early (P3/P4) and late (P10) passages were collected for the in vitro analyses including: the sequencing of genes frequently mutated in OS and EWS, evaluation of telomerase activity, assessment of the gene expression profile and activity of major cancer pathways, cytogenetic analysis on synchronous MSC, and molecular karyotyping using a comparative genomic hybridization (CGH) array.
MSC-SAR displayed comparable morphology, immunophenotype, proliferation rate, differentiation potential, and telomerase activity to MSC-CTRL. Both cell types displayed signs of senescence in the late stages of culture with no relevant changes in cancer gene expression. However, cytogenetic analysis detected chromosomal anomalies in the early and late stages of MSC-SAR and MSC-CTRL after culture.
Our results demonstrated that the in vitro expansion of MSC does not influence or favor malignant transformation since MSC-SAR were not more prone than MSC-CTRL to deleterious changes during culture. However, the presence of chromosomal aberrations supports rigorous phenotypic, functional and genetic evaluation of the biosafety of MSCs, which is important for clinical applications.
Full-text · Article · Apr 2014 · Journal of Translational Medicine
[Show abstract][Hide abstract] ABSTRACT: The umbilical cord (UC) is a promising source of mesenchymal stromal cells (MSCs). UC-MSCs display very similar in vitro characteristics to bone marrow-MSCs and could represent a valuable alternative for cell-based therapies. However, it is still unclear whether UC-MSCs are prone or not to the acquisition of genomic imbalances during in vitro expansion.
With the use of array-comparative genomic hybridization, we compared copy number variations of early (P2-P3) and late (>P5) passages of in vitro-expanded UC-MSCs.
In two of 11 long-term UC-MSCs cultures, we observed the appearance of clones carrying genomic imbalances, which generated genetic mosaicism at intermediate passages. Although still able to reach the senescence phase, the cells carrying the genomic imbalance acquired a proliferative advantage, as demonstrated by the increase in frequency during long-term culture.
Altogether, our results suggest that UC-MSC-based clinical protocols should be designed with caution; their clinical use should be preceded by array-comparative genomic hybridization screening for the acquisition of genomic imbalances during in vitro expansion.
[Show abstract][Hide abstract] ABSTRACT: We report the first case of renal antineutrophil cytoplasmic antibody (ANCA)-associated vasculitis treated with autologous mesenchymal stromal cells (MSCs). A 73-year-old man was admitted to the hospital for malaise, weight loss, and oliguria. His serum creatinine level was 2.7 mg/dL but it rapidly increased to 7.8 mg/dL; urinalysis showed proteinuria and hematuria, and the ANCA to myeloperoxidase with a perinuclear pattern (pANCA) titer was high (132 IU/mL). Renal biopsy showed necrotizing crescentic glomerulonephritis. Standard immunosuppressive therapy (cyclophosphamide and corticosteroids) was ineffective. Rituximab therapy was started, but it was discontinued after the third dose to minimize the risk of systemic spread of a severe oral Candida infection and to prevent superinfections that were facilitated by leukopenia. The patient received autologous MSCs, 1.5 × 10(6) cells/kg body weight, intravenously. After 7 days, his serum creatinine level decreased to 2.2 mg/dL, pANCA titer decreased to 75 IU/mL, and urinalysis findings normalized. Eight months later, he received a second MSC infusion because his serum creatinine level increased. In 1 week, his creatinine level decreased to 1.9 mg/dL and his pANCA titer decreased to 14 IU/mL. Immunosuppressive therapy was subsequently withdrawn. At the last follow-up visit, 12 months after the second MSC infusion, the patient remained in clinical remission without any therapy. Infusion of MSCs induced expansion of the T-lymphocyte subset expressing a regulatory T-cell phenotype (CD4(+)CD25(+)Foxp3(+)) and a notable reduction in interferon-γ, interleukin 6, and tumor necrosis factor serum levels.
Full-text · Article · Oct 2013 · Mayo Clinic Proceedings
[Show abstract][Hide abstract] ABSTRACT: Alterations in hematopoietic microenvironment of acute lymphoblastic leukemia patients have been claimed to occur, but little is known about the components of marrow stroma in these patients. In this study, we characterized mesenchymal stromal cells (MSCs) isolated from bone marrow (BM) of 45 pediatric patients with acute lymphoblastic leukemia (ALL-MSCs) at diagnosis (day+0) and during chemotherapy treatment (days: +15; +33; +78), the time points being chosen according to the schedule of BM aspirates required by the AIEOP-BFM ALL 2009 treatment protocol. Morphology, proliferative capacity, immunophenotype, differentiation potential, immunomodulatory properties and ability to support long-term hematopoiesis of ALL-MSCs were analysed and compared with those from 41 healthy donors (HD-MSCs). ALL-MSCs were also genetically characterized through array-CGH, conventional karyotyping and FISH analysis. Moreover, we compared ALL-MSCs generated at day+0 with those isolated during chemotherapy. Morphology, immunophenotype, differentiation potential and in vitro life-span did not differ between ALL-MSCs and HD-MSCs. ALL-MSCs showed significantly lower proliferative capacity (p<0.001) and ability to support in vitro hematopoiesis (p = 0.04) as compared with HD-MSCs, while they had similar capacity to inhibit in vitro mitogen-induced T-cell proliferation (p = N.S.). ALL-MSCs showed neither the typical translocations carried by the leukemic clone (when present), nor other genetic abnormalities acquired during ex vivo culture. Our findings indicate that ALL-MSCs display reduced ability to proliferate and to support long-term hematopoiesis in vitro. ALL-MSCs isolated at diagnosis do not differ from those obtained during treatment.
[Show abstract][Hide abstract] ABSTRACT: In the present study, gelatin-based cryogels have been seeded with human SAOS-2 osteoblasts. In order to overcome the drawbacks associated with in vitro culture systems, such as limited diffusion and inhomogeneous cell-matrix distribution, this work describes the application of ultrasounds (average power, 149 mW; frequency, 1.5 MHz) to physically enhance the cell culture in vitro. The results indicate that the physical stimulation of cell-seeded gelatin-based cryogels upregulates the bone matrix production.
No preview · Article · Jul 2013 · Conference proceedings: ... Annual International Conference of the IEEE Engineering in Medicine and Biology Society. IEEE Engineering in Medicine and Biology Society. Conference
[Show abstract][Hide abstract] ABSTRACT: Some biological properties of Bombyx mori sericins from twenty strains were investigated, fourteen fed with artificial diet, two with fresh mulberry leaves and four with both diets. Sericin exhibited ROS-scavenging, anti-tyrosinase and anti-elastase properties, the strain significantly influencedthese properties, while diet only influenced the anti-tyrosinase activity. Sericins were clustered into 5 groups and one sericin from each group was further studied: sericins showed anti-proliferative activity on in vitro stimulated peripheral blood mononuclear cells; some strains decreased in vitro secretion of IFNγ, while no effects were observed on TNFα and IL10 release. Therefore, a mixture of sericins extracted from the most promising strains may be useful for dermatological and cosmetic use.
Full-text · Article · Mar 2013 · International journal of biological macromolecules
[Show abstract][Hide abstract] ABSTRACT: Mesenchymal stem cells (MSCs) were first isolated more than 50 years ago from the bone marrow. Currently MSCs may also be isolated from several alternative sources and they have been used in more than a hundred clinical trials worldwide to treat a wide variety of diseases. The MSCs mechanism of action is undefined and currently under investigation. For in vivo purposes MSCs must be produced in compliance with good manufacturing practices and this has stimulated research on MSCs characterization and safety. The objective of this review is to describe recent developments regarding MSCs properties, physiological effects, delivery, clinical applications and possible side effects.
Full-text · Article · Dec 2012 · Current pharmaceutical design
[Show abstract][Hide abstract] ABSTRACT: Adult autoimmune enteropathy (AIE) is a rare cause of malabsorption syndrome unresponsive to dietary restriction. Its diagnostic hallmarks are small-bowel villous atrophy and antienterocyte autoantibodies. Therapy is based mainly on nutritional support and immunosuppression. We treated a 61-year-old woman with corticosteroid-refractory AIE and life-threatening malabsorption syndrome with systemic infusions of autologous, bone marrow-derived, mesenchymal stromal cells (MSCs) as rescue therapy. The MSCs were expanded ex vivo following a previously used Good Manufacturing Practice procedure, and 2 intravenous infusions of 1.8 × 10(6) MSCs/kg body weight were administered 2 weeks apart. Analysis of circulating and mucosal regulatory T-and B-cell numbers, and of serum and secretory immunoglobulin levels, was performed before and after treatment. The MSC infusions were safe and effective, leading to disappearance of disease hallmarks and recovery from the life-threatening condition. Increases in mucosal regulatory T-cell numbers and secretory immunoglobulin levels were also observed. The benefit, however, was transient, and a further MSC infusion resulted in the same short efficacy. This case encourages the use of MSCs to treat patients with life-threatening, corticosteroid-refractory AIE and suggests that MSC infusion can attenuate, albeit transiently, the autoimmune attack.
No preview · Article · Sep 2012 · Mayo Clinic Proceedings