Treatment of pseudoarthrosis of the upper limb using expanded mesenchymal stem cells: a pilot study.

BACKGROUND: In orthopedic field is growing interest in the use of stem cells: this mesenchymal multipotent line (MSCs) can lead to differentiation into osteocytes and thus the formation of bone tissue. In literature applications of this line are described in injuries of tendons and ligaments, small bony avulsions, nonunion fractures and cartilage defects. AIM: Utilize MSCs expanded in laboratory in case of atrophic pseudoarthrosis of the upper limb. MATERIALS AND METHODS: We obtain the amount of cell necessary for the implant by the collaboration with the UO Haematological Department. For the procedure we make a blood sample from the iliac crest bone marrow and a subsequent phase of selection and cultivation of mesenchymal line for 3 weeks, to get a sufficient amount of tissue to be used, which is presented at the time of surgery on a scaffold made by autologous plasma gel and CaCl(2). We reassessed our experience in 8 different types of upper limb fractures result in pseudarthrosis and delayed of consolidation: 4 women and 4 men, average 44 years old followed with a follow-up of 50.3 months. In all cases the site of non-union has been revitalized (by microfractures and drilling) and a synthesis was performed with a rigid plate. So we fill the bone gap with autologous bone and mesenchymal stem cells expanded in the laboratory. RESULTS: We have a radiographic healing in 8 cases and no adverse events were highlighted. CONCLUSIONS: Using this cells line we obtained encouraging but certainly not conclusive impressions, according to the limited number of cases and lack of adequate comparative studies. In tissue engineering are also certainly needed further investigations and developments.

Mesenchymal cells inhibit expansion but not cytotoxicity exerted by gamma-delta T cells.

BACKGROUND: Multipotent mesenchymal stromal cells (MSCs) exert a relevant immunosuppressive activity by inhibiting T- and B-lymphocytes, natural killer (NK) cells and dendritic cell expansion. Nevertheless, a possible activity on gamma/delta T cells has still not been evaluated. Gamma-delta T lymphocytes play an important role in the control of cancer and they have been shown to be implicated in graft-vs.-host disease. Thus, modulation of activation and proliferation of these cells could be relevant for therapeutic purposes. MATERIALS AND METHODS: Peripheral blood mononuclear cells from 21 healthy donors were used as source for gamma-delta T cells, expanded in presence of 10 IU mL(-1) interleukin-2 (IL-2) and 1 microM zoledronate. MSCs were recovered from patients undergoing routine total hip replacement surgery, and characterised by flow cytometry. Cytotoxicity on multiple myeloma and melanoma cell lines was assessed by measuring dilution of the carboxyfluorescein diacetate succinimydylester dye (CFSE). Gamma-delta T cells were then incubated with MSCs in contact cultures, and with addition of MSC-conditioned medium. RESULTS: In this article we confirmed that (1) in vitro expanded gamma-delta T cells play a significant anti-proliferative effect on multiple myeloma and melanoma cells and (2) multipotent mesenchymal stromal cells effectively suppress the ex vivo expansion of T cells carrying a specific T-cell receptor gene (TCR) rearrangement, Vgamma9/Vdelta2, induced by the combination of IL-2 and zoledronate, without interfering with their cytotoxic activity. DISCUSSION: These findings contribute to explain the activity of ex vivo expanded mesenchymal cells, suggesting that MSCs would interact with gamma-delta T lymphocytes. CONCLUSION: This effect could be relevant in separating graft-vs.-host from the graft-vs.-tumour effect, especially considering the possibility of modulating T-lymphocytes activity by the immunomodulating drugs now available.

PS-341 (Bortezomib) inhibits proliferation and induces apoptosis of megakaryoblastic MO7-e cells.

PS-341 (Bortezomib) is a dipeptide boronic acid proteasome inhibitor with antitumor activity that induces apoptosis in different human cancer cell lines. We investigated effects of PS-341 (Bortezomib) on cell proliferation, cell cycle progression, induction of apoptosis and differentiation in a megakaryoblastic (MO7-e) cell line. PS-341 was able to retain NF-kappaB in the cytoplasm and inhibit cell growth (IC(50)=22.5 nM), in a dose/time-dependent way. This anti-proliferative activity resulted to be lineage-specific, because other leukemic cell lines (KG1a, K562/R7, HL60/DNR) were unaffected by the PS-341 treatment. Moreover, PS-341 in MO7-e induced a significant pro-apoptotic effect from 10 nM concentration (40% versus 12% in the control, p<0.05). On the other hand, at lower concentration (5 nM), Bortezomib blocked cell cycle in the G2 phase. Finally, this compound was able to down-regulate WT1 expression. No significant effects on cell differentiation were found. Because a spontaneous NF-kappaB activation has been reported in megakaryocytes from patients affected by myeloproliferative disorders, Bortezomib would so be an attractive therapeutic tool for these malignancies, including essential thrombocythemia or idiopathic myelofibrosis. Preliminary data show an inhibiting activity of Bortezomib in the megakaryocytic colonies formation. Finally, also down-regulation of the WT1 gene Bortezomib-driven could be relevant, because of the role that this gene would play in the pathogenesis of acute and chronic myeloproliferative disorders.

The efficacy of rituximab plus Hyper-CVAD regimen in mantle cell lymphoma is independent of FCgammaRIIIa and FCgammaRIIa polymorphisms.

Mantle cell lymphoma (MCL) accounts for 3-10% of all non-Hodgkin's lymphomas, with median overall survival not exceeding 3-4 years. Rituximab in combination with the Hyper-CVAD regimen appears the most promising regimen; thus, we adopted it as a first-line treatment strategy in a series of 24 patients. In addition to evaluation of clinical success of the regimen, we investigated a possible role of polymorphism in IgG Fc receptors, FCgammaRIIIa and FCgammaRIIa. The frequencies of FCgammaRIIIa-158 were as follows: V/V=4/24 (17%); V/F=16/24 (66%); F/F=4/24 (17%). Those of the FCgammaRIIa-131 polymorphism were H/H=11/24 (46%), H/R=9/24 (37%), R/R=4/24 (17%). The overall response rate was 62.5%, with 33% of complete responses (CRs) after four cycles of R-Hyper-CVAD. Two-year progression-free survival (PFS) was 78% for 158V/V patients vs 75% for cases carrying phenylalanine (p=0.88). When the FCgammaRIIa polymorphism was assessed, the 2-year PFS was 82% for 131H/H patients vs 75% for those carrying arginine (p=0.26). Eighty-three percent of cases achieved Polymerase Chain Reaction (PCR)-negativity: the progression rate was significantly influenced by the minimal residual disease clearance, with 12% progression in the subgroup of PCR-negative cases versus 67% progression in PCR-positive cases (p=0.008). The achievement of PCRnegativity was not significantly influenced by FCgammaR polymorphisms. Results confirm that rituximab plus Hyper-CVAD is an effective regimen for the induction of prolonged remission in patients with aggressive MCL and suggest that rituximab efficacy is independent of the FCgammaR polymorphisms.

Contemporaneous appearance, 18 years after allogeneic bone marrow transplantation, of myelodysplastic syndrome in the patient and the donor.

We report the case of the development of two different stages of the same clonal disorder in two patients sharing the same bone marrow due to a previous bone marrow allotransplant. The transplanted patient developed severe aplasia with myeloid blasts, different from those of the previously cured leukemia. Chimerism evaluated by microsatellite analyses confirmed a full donor phenotype. At the same time, the donor of the bone marrow transplantation developed a refractory anemia with excess blasts. We speculate on the presence of an undetectable pre-existing pathological clone in the transplanted bone marrow, which have evolved in the two patients.

A myeloablative allograft after rejection of two consecutive nonmyeloablative transplants from two different HLA identical siblings.

A 55-year-old female with standard risk AML in second CR received an allogenic transplant from an HLA-matched sibling, using a nonmyeloablative conditioning regimen (NMST). On day +139, she rejected her graft with autologous reconstitution. She received a second NMST from a different HLA-matched sibling with an identical conditioning regimen and immunosuppression. On day +110, she rejected the second graft, with autologous reconstitution with blasts. She received a third allograft from the first sibling with a myeloablative busulfan-based conditioning regimen. She is now day +270, in CR, with full donor chimerism.

Arsenic and all-trans retinoic acid as induction therapy before autograft in a case of relapsed resistant secondary acute promyelocytic leukemia.

Arsenic trioxide has recently been reported to be successful in the treatment of promyelocytic leukemia. Several concerns about the use of this toxic agent are currently reducing its potential clinical use even in severely ill patients. In this report we describe the results achieved by As2O3 with all-trans retinoic acid in a patient suffering from secondary, relapsed, resistant promyelocytic leukemia. Several complications, including sepsis and an extensive area of skin necrosis, did not allow us to treat the patient further with chemotherapy. With As2O3 and ATRA therapy, the patient obtained a complete molecular remission without any significant toxicity and, subsequently, it was possible to perform a bone marrow autograft in a state of complete remission.

Quantitative molecular evaluation in autotransplant programs for follicular lymphoma: efficacy of in vivo purging by Rituximab.

The main aim of this paper was to compare results of Genescan and real-time PCR methods in order to detect contamination in harvests from patients with follicular lymphoma. The secondary goal was to evaluate the efficacy of Rituximab as an in vivo purging agent. A total of 23 patients had been treated with CHOP followed by either high-dose therapy (12 patients) or high-dose plus Rituximab (11 patients), both followed by autologous transplantation. Results show that 86% of harvests from patients treated with Rituximab were PCR-negative compared to 14.3% from controls. Real-time PCR was more sensitive than Genescan PCR; quantitative analysis revealed a correlation between the amount of contamination in the harvests and relapse after transplantation. Whereas all patients reinfused with negative aphereses achieved complete remission and showed a significantly better 5-year PFS (100%) compared to those reinfused with contaminated samples (41%), a very low amount of contamination does not appear to negatively affect outcome, suggesting that determination of a cutoff in the contamination level of harvests could be useful. Results suggest that real-time PCR is superior to Genescan PCR to select transplantable harvests and confirm the ability of Rituximab as an in vivo purging tool for follicular lymphoma.

The role of molecular monitoring in autotransplantation for non-Hodgkin's lymphoma.

Seventy-two patients with non-Hodgkin's lymphoma were evaluated for the presence of molecular markers (IgH, bcl-1, bcl-2 rearrangement) on bone marrow, at diagnosis and after PBSCT, and on harvests in order to find a possible predictive role of minimal residual disease on treatment outcome. At diagnosis, 41 (59%) out of 69 available bone marrows showed molecular involvement. Fifty-six percent of leukaphereses were involved, mainly indolent lymphoma (P = 0.001) or advanced disease (P = 0.01). Ex vivo purging cleared only one stem collection out of 31 PCR-positive leukaphereses. Aggressive lymphomas showed both a longer overall survival (OS) (P = 0.03) and relapse-free survival RFS (P = 0.02) when transplanted with unpurged stem cells, whereas indolent NHL survival was not influenced by ex vivo purging. Twenty out of 26 samples taken during follow-up had bone marrow involvement at diagnosis. Of these, 15 cleared their bone marrow; both OS and RFS were significantly longer in the PCR-negative cases (P = 0.05 and P = 0.005). At 1 year after PBSCT, 75% of patients were PCR negative, with 50% molecular remissions; the relapse rate was 55% for patients still PCR positive vs 29% for those who were PCR negative. Thus, after high-dose chemotherapy, close molecular monitoring of MRD using qualitative PCR techniques seems to represent a reliable prognostic indicator.

Oral cyclophosphamide therapy for patients with residual or relapsed indolent-type lymphoma after initial treatment for aggressive lymphomas. A sub-group of patients with apparent transformed indolent lymphoma.

Lymph node or bone marrow biopsy from sixty-one patients affected by aggressive non-Hodgkin lymphomas (NHL) were retrospectively evaluated to assess the histology at relapse. Eighteen cases (29.5%) were proven to have relapsed or persistent low-grade lymphoma after conventional therapy. In 5/18 patients association of low and high-grade lymphoma was detectable at diagnosis by bone marrow biopsy. In the remaining 13/18 no evidence of follicular lymphoma was detected at diagnosis. The outcome of these patients was compared to that of 43 patients relapsed without change in histology and treated by a second line therapy. Of these 43 patients, 13 were not responders (NR), 10 achieved a partial remission (PR) and 18 complete remission (CR). Two were lost during follow-up. The 18 patients with residual/relapsed indolent subtype received oral cyclophosphamide (100 mg/day for 15 days every month for six months): 3 of them had NR, 5 CR, and 10 PR. The overall survival (OS) median time was 39 months in low-grade resistant/relapsed patients and 20 months in patients with aggressive histology. OS at 24 months was 71 and 41%, respectively, (p < 0.02). Most of the patients with high-grade disease were refractory or relapsed after a median of five months, whereas cases with low-grade NHL showed a long lasting stable PR. We suggest that the higher grade patients with residual or relapsed low grade lymphoma were, in fact, transformed low-grade at diagnosis and, after removing the more aggressive component by chemotherapy, it is possible to manage these patients by conventional therapy for indolent lymphomas.

Early reappearance of primary solid cancer in patients treated with purine analogs.

Two patients, observed at our institution, developed, after treatment with fludarabine, an early reappearance of metastatic primary solid cancers which were previously in long-lasting, complete remission. Patients had earlier suffered from a solid cancer considered cured and, subsequently, developed a lymphoid disorder treated with fludarabine. The two patients developed histologically confirmed hepatic metastasis from breast cancer and colic adenocarcinoma respectively 11 and 4 months after the beginning of fludarabine-therapy. Purine analogs have been reported to be effective against chronic lymphocytic leukemia and indolent lymphomas. However, these drugs induce severe immunodeficiency. In addition to the infectious diseases related to the treatment, the use of these drugs could facilitate the development of secondary neoplasms, related to the patient's impaired immunosurveillance. The surprisingly short latency between the therapy and the reappearance of non hematological cancers seen in our patients suggests that treatment with purine analogs may be involved in the reappearance of the tumors. In this regard, we suggest a possible role for purine analog-induced immunodeficiency in allowing the growth of previously undetected cancer cells rather than a direct drug-related mutagenic activity.

The clinical relevance of the expression of several multidrug-resistant-related genes in patients with primary acute myeloid leukemia.

Multidrug resistance (MDR) is a complex phenomenon that includes the expression of many different genes regulating drug transport or metabolism, cellular repair or detoxification mechanisms. The co-expression of several genes could be at the basis of the resistant phenotype in vivo. In order to test a possible prognostic role of the expression and co-expression of several MDR-related genes (MDR1, topoisomerase IIalpha, topoisomerase IIbeta, MRP, GSTpi, LRP), 35 patients affected by acute myeloid leukemia (AML) were tested by RT-PCR assays. In our series, topoisomerase IIbeta was significantly co-expressed with MRP (p = 0.05), GSTpi (p = 0.017) and LRP (p = 0.005). GSTpi was co-expressed with LRP (p = 0.03) and MRP (p = 0.007); on the other hand, 53.8% of patients were LRP and MRP-positive (p = 0.02). The PCR-positivity did not differ according to biological/clinical characteristics of patients, including age; this latter was the only parameter conditioning the response and overall survival. Neither the expression nor the co-expression of the tested genes was significantly correlated with the response to the induction treatment and long-term outcome.

Laparoscopic diagnosis of tubaric pregnancy in evolution and reconstructive surgical treatment.

The authors report the results obtained on 22 selected cases of tubaric pregnancy in evolution and treated by reconstructive plastic surgery. They point out the necessity of a very precocious laparoscopic diagnosis in order to obtain a better functional recovery of the fallopian tube. On the basis of the results, the authors suggest a more frequent reconstructive treatment of the tubaric pregnancy.

Balancing books & balancing values: the hidden threat of PPS.

Losing sight of the real vision and values of home care is a threat that becomes more real as agency leaders try to redefine their agencies to be PPS ready. Agencies must remain committed to ensuring that the foundation of the organization is one of sound business practices coupled with solid core values.

Exceptional services, exceptional caring.

Responding professionally and compassionately to the health care needs of clients has always been the number-one priority of home care agencies. Recently, however, agencies have become aware of the significant implications that customer service has on the financial future of their organizations.

The competitive edge of dynamic marketing.

Home care is becoming a more and more competitive field. The key to success in competition is differentiating yourself from the competitors. And the key to that is exceptional customer service--going above and beyond what your customer expects.

A new millennium .... a new home care.

Home care faces many changes--from funding to changing patient populations. Agencies need to assess what they are doing and build into their missions services that take into account these changes. How can they do that?

What home care executives should know about managed care organizations: preliminary results from a national study.

An industry survey explored the growing relationship home care has with managed care by asking managed care organizations (MCOs) what they know about home care, what they want from home care, and what they expect from home care agencies. The preliminary results here should help home care providers develop a working partnership with MCOs that will be successful now and in the future.

Collaborating to compete: a national study of horizontal networks.

Horizontal networks offer providers challenges as well as options. To help providers explore the idea of joining a network, a national study presents the experiences of successful and unsuccessful home care networks.