Adipose-derived regenerative cells and fat grafting for treating breast cancer-related lymphedema: Lymphoscintigraphic evaluation with 1 year of follow-up.

BACKGROUND: Breast cancer-related lymphedema (BCRL) is a feared late complication. Treatment options are lacking at present. Recent studies have suggested that mesenchymal stromal cells can alleviate lymphedema. Herein, we report the results from the first human pilot study with adipose-derived regenerative cells (ADRCs) for treating BCRL with 1 year of follow-up. MATERIAL AND METHODS: We included 10 patients with BCRL. ADRCs were injected directly into the axillary region together with a scar-releasing fat grafting procedure. Primary endpoint was change in arm volume. Secondary endpoints were change in patient-reported outcomes, changes in lymph flow, and safety. RESULTS: During follow-up, no significant change in volume was noted. Patient-reported outcomes improved significantly with time. Five patients reduced their use of conservative management. Quantitative lymphoscintigraphy did not improve on the lymphedema-affected arms. ADRCs were well tolerated, and only minor transient adverse events related to liposuction were noted. CONCLUSIONS: In this pilot study, a single injection of ADRCs improved lymphedema based on patient-reported outcome measures, and there were no serious adverse events during the follow-up period. Lymphoscintigraphic evaluation showed no improvement after ADRC treatment. There was no change in excess arm volume. Results of this trial need to be confirmed in randomized clinical trials.

Human serum levels of fetal antigen 1 (FA1/Dlk1) increase with obesity, are negatively associated with insulin sensitivity and modulate inflammation in vitro.

OBJECTIVE: To investigate fetal antigen 1 (FA1) protein within the context of human obesity and its relation with insulin sensitivity. SUBJECTS: Cross-sectional study that analyses circulating levels of FA1 in two selected human cohorts: n=127 men for the study of FA1 circulating levels in the context of obesity and insulin sensitivity (S(i)); and n=61 severely obese women before and after bariatric surgery. The response in vitro to FA1 protein on human cell lines of monocytes, preadipocytes and mature adipocytes was studied. MEASUREMENTS: Anthropometrical parameters: body mass index, waist-to-hip ratio, waist circumference, fat-free mass and fat mass. Clinical parameters: lipid profile (high-density lipoprotein (HDL) cholesterol, low-density lipoprotein (LDL) cholesterol, total cholesterol, triglycerides), glycemic profile (fasting glucose, insulin, S(i), HOMA-IR (Homeostasis Model Assessment of Insulin Resistance), cytokines (sIL-6), adipokines (adiponectin) and circulating soluble fractions of tumor necrosis factor-alpha receptors 1 and 2 (sTNFR1 and sTNFR2). RESULTS: IN the obesity study, levels of FA1 in serum were found to increase with obesity. The S(i) index was negatively dependent on FA1 levels. In severe obesity, serum levels of FA1 decreased 1.4-fold 6 months after bariatric surgery. In vitro assays with FA1 protein on human monocytes and adipocytes cell lines modified the expression of pro-inflammatory cytokines and adipokines (tumor necrosis factor-alpha (TNFalpha), monocyte chemoattractant protein-1 (MCP-1), IL-6 (interleukin-6) and adiponectin). CONCLUSION: FA1 serum levels were increased in obese subjects and might influence S(i). The stimulatory effect of FA1 protein on pro-inflammatory cytokines on both immune and adipose cell types could contribute to worsening the inflammatory environment observed in obesity.

Dlk1 in normal and abnormal hematopoiesis.

Dlk1 (Pref-1) is a transmembrane and secreted protein, which is a member of the epidermal growth factor-like family, homologous to Notch/Delta/Serrate. We have found by real-time RT-PCR that Dlk1 mRNA levels were high in CD34(+) cells in 10 of 12 MDS samples compared with CD34(+) cells from 11 normals. Also, Dlk1 mRNA was elevated in mononuclear, low density bone marrow cells from 11/38 MDS patients, 5/11 AML M6 and 2/4 AML M7 samples. Furthermore, 5/6 erythroleukemia and 2/2 megakaryocytic leukemia cell lines highly expressed Dlk1 mRNA. Levels of Dlk1 mRNA markedly increased during megakaryocytic differentiation of both CMK megakaryoblasts as well as normal CD34(+) hematopoietic stem cells. High serum levels of Dlk1 occurred in RA (4/10) and essential thrombocythemia (2/10) patients. Functional studies showed that forced expression of Dlk1 enhanced proliferation of K562 cells growing in 1% fetal bovine serum. Analysis of hematopoiesis of Dlk1 knockout mice suggested that Dlk1 contributed to granulocyte, megakaryocyte and B-cell clonogenic growth and was needed for generation of splenic B-cells. In summary, Dlk1 is overexpressed in selected samples of MDS (especially RA and RAEB) and AML (particularly M6, M7), and it appears to be associated with normal development of megakaryocytes and B cells.

Carrier screening for Canavan disease in Australia.

This study reports, for the first time, the carrier frequency of Canavan disease in the Ashkenazi Jewish population in Australia, and the identification of a novel mutation in the ASPA gene.

Expression, biosynthesis and release of preadipocyte factor-1/ delta-like protein/fetal antigen-1 in pancreatic beta-cells: possible physiological implications.

Preadipocyte factor-1 (Pref-1)/delta-like protein/fetal antigen-1 (FA1) is a member of the epidermal growth factor-like family. It is widely expressed in embryonic tissues, whereas in adults it is confined to the adrenal gland, the anterior pituitary, the endocrine pancreas, the testis and the ovaries. We have previously cloned Pref-1 from neonatal rat islets stimulated by GH. The aim of the present study was to elucidate the biosynthesis and release of Pref-1/FA1 in beta-cells and to determine if Pref-1/FA1 is mediating the mitogenic effect of GH in insulin-producing cells. First we studied the biosynthesis and processing of Pref-1 to the soluble form, FA1, in pancreatic islets and insulinoma cells transfected with Pref-1 cDNA. We measured the release of FA1 by ELISA and the possible effect of FA1 in GH-stimulated beta-cell proliferation by incorporation of bromodeoxyuridine (BrdU) in insulin-positive islet cells. We found that Pref-1 was synthesized in normal islets and in RINm5F insulinoma cells and released into the medium in two forms, of which one corresponded to FA1. Both the expression of the mRNA for Pref-1 and the release of the soluble form(s) were stimulated by GH and prolactin (PRL). Whereas 2 h exposure to high glucose or 3-isobutyl-1-methylxanthine stimulated insulin release, only a small change was seen in FA1 release, suggesting that the FA1 is released by a different pathway than insulin. However, long-term exposure (48 h) to high glucose increased FA1 secretion, indicating that FA1 is regulated by glucose. Neither FA1 nor conditioned medium from GH-stimulated islets depleted for GH was able to increase beta-cell replication and overexpression of Pref-1 resulted in attenuated proliferation of the RINm5F cells. By immunocytochemistry of GH-stimulated islet cells no correlation between high Pref-1 expression and BrdU incorporation was observed and there was an inverse relationship between the levels of insulin and Pref-1. These results indicate that Pref-1/FA1 is not mediating the mitogenic effect of GH and PRL. Therefore the function of Pref-1 in the beta-cell remains unknown.

Neurons in the monoaminergic nuclei of the rat and human central nervous system express FA1/dlk.

The gene DLK1 encodes a member of the epidermal growth factor (EGF) superfamily, delta-like (dlk). When exposed in vivo to the action of an unknown protease, this type 1 membrane protein generates a soluble peptide referred to as Fetal antigen 1 (FA1). By acting in juxtacrine as well as paracrine/autocrine manners, both forms have been shown to be active in the differentiation/proliferation process of various cell types. In adults, FA1/dlk has been demonstrated mainly within (neuro) endocrine tissues. In this study we investigated the presence of FA1/dlk in other parts of the developing and adult rat and human CNS. Using immunocytochemistry and in situ hybridization we found that in both species FA1/dlk was expressed in neurons of the Edinger-Westphal's nucleus as well as in substantia nigra, ventral tegmental area (VTA), locus coeruleus and in certain parts of the raphe nuclei.

Fetal antigen 1 in healthy adults and patients with pituitary disease: relation to physiological, pathological, and pharmacological GH levels.

Immunohistochemical analysis of the distribution of human fetal antigen 1 (FA1) in adult human tissues has demonstrated a strong association between FA1 and (neuro)endocrine structures. In the anterior pituitary gland FA1 was colocalized with GH, and the present study was performed to evaluate a possible relationship between GH and FA1. FA1 and GH levels were measured during a 24-h period at 20-min intervals. In contrast to the known GH peaks during 24-h sampling, there was no detectable FA1 peak. The FA1 responses to placebo were not significantly different from the responses to the combination of pyridostigmine and GHRH. No significant difference was found between basal FA1 (nanograms per ml) levels [median (minimum-maximum)] in healthy adults [n = 40; 28.6 ng/ml (12.5-72.0)], acromegalic patients [n = 11; 31.0 ng/ml (21.6-56.3)], and patients with GH deficiency [n = 22; 32.1 ng/ml (13.4-108.7)]. FA1 levels were significantly reduced, in the six of seven acromegalic GH responders to octreotide, from [median (minimum-maximum)] 30.6 ng/ml (20.0-43.1) to 20.3 (13.9-30.2; P < 0.02). There was no significant change during placebo. FA1 levels were significantly increased compared with placebo values during 3 months of GH therapy. The increase in FA1 levels was significantly higher than the change during placebo (P < 0.003). In conclusion, a common secretory and stimulatory pathway for FA1 and GH in healthy adults has been ruled out. However, we found that pharmacologically induced changes in GH levels during weeks to months had a corresponding direct or indirect effect on FA1 levels in patients with GH deficiency or acromegaly. However, a direct effect of octreotide on FA1 levels, independent of GH levels, has not been ruled out.

Quantitative magnetic resonance imaging as marker of synovial membrane regeneration and recurrence of synovitis after arthroscopic knee joint synovectomy: a one year follow up study.

OBJECTIVES: By repeated magnetic resonance imaging (MRI) to study synovial membrane regeneration and recurrence of synovitis after arthroscopic knee joint synovectomy in patients with rheumatoid arthritis (RA) and other (non-RA) causes of persistent knee joint synovitis. METHODS: Contrast enhanced MRI was performed in 15 knees (nine RA, six non-RA) before and one day, seven days, two months, and 12 months after arthroscopic synovectomy. Synovial membrane volumes, joint effusion volumes, and cartilage and bone destruction were assessed on each MRI set. Baseline microscopic and macroscopic assessments of synovitis and baseline and follow up standard clinical and biochemical examinations were available. RESULTS: Synovial membrane and joint fluid volumes were significantly reduced two and 12 months after synovectomy. However, MRI signs of recurrent synovitis were already present in most knees at two months. No significant differences between volumes in RA and non-RA knees were seen. Synovial membrane volumes at two months were significantly inversely correlated with the duration of clinical remission, for all knees considered together (Spearman's correlation r(s)=-0.67; p<0.05), for RA knees (r(s)=-0.76; p<0.05), and for non-RA knees (r(s)=-0.83; p<0.05). Baseline volumes were not significantly correlated with clinical outcome. Only three knees (all RA) showed erosive progression. The rate of erosive progression was not correlated with MRI volumes or with clinical or biochemical parameters. CONCLUSION: The synovial membrane had regenerated two months after arthroscopic knee joint synovectomy and despite significant volume reductions compared with baseline it often showed signs of recurrent synovitis. MRI seems to be valuable as a marker of inflammation, destruction and, perhaps, as a predictor of therapeutic outcome in arthritis.

Flow cytometric detection of growth factor receptors in autografts and analysis of growth factor concentrations in autologous stem cell transplantation: possible significance for platelet recovery.

In order to improve prediction of hematopoietic recovery, we conducted a pilot study, analyzing the significance of growth factor receptor expression in autografts as well as endogenous growth factor levels in blood before, during and after stem cell transplantation. Three early acting (stem cell factor (SCF), Flt3 ligand (Flt3) and fetal antigen 1 (FA1)) and three lineage-specific growth factors (EPO, G-CSF and thrombopoietin (Tpo)) were analyzed by ELISA in 16 patients with multiple myeloma (MM) and 16 patients with non-Hodgkin's lymphoma (NHL). The relative number of SCF, Flt3, Tpo and G-CSF receptor positive, CD34+ progenitor cells were measured by flow cytometry in the leukapheresis product used for transplantation in a subgroup of 15 patients (NHL, n = 8, MM, n = 7). Three factors were identified as having a significant impact on platelet recovery. First, the level of Tpo in blood at the time of the nadir (day +7). Second, the percentage of re-infused thrombopoietin receptor positive progenitors and finally, the percentage of Flt3 receptor positive progenitors. On the other hand, none of the analyzed factors significantly predicted myeloid or erythroid recovery. These findings need to be confirmed in prospectively designed studies.

Follow-up of volar plate interposition arthroplasty (Tupper) of the metacarpophalangeal joints in rheumatoid hands: preliminary findings.

We present the results of a three-year follow-up study of 59 interposition volar plate arthroplasties (Tupper) on metacarpophalangeal joints in 13 patients with erosive rheumatoid arthritis. The median age at the time of operation was 60 years (range 45-77). All patients reported lasting pain relief at rest. Improvement of hand function was, however, less satisfactory as both grip and pinch strength were compromised in all patients. Seven patients were satisfied with the outcome whereas six complained of stiffness in the operated joints. All but one patient concluded that they would have the operation again if necessary.

Weilby-Burton arthroplasty of the trapeziometacarpal joint of the thumb.

Twenty-three patients (25 thumbs) were treated by tendon interposition arthroplasty for trapeziometacarpal arthrosis as described by Weilby and modified slightly as described by Burton and Pellegrini. There was good (4/25, 16%) or complete (19/25, 76%) pain relief in 23 (92%) of the cases. Activities of daily living were generally easier. Mobility and strength of the thumb were satisfactory. One patient had signs of instability during a stress test. We conclude that our technique produces a stable and pain-free thumb joint. However, careful selection of the patients for this procedure is essential, and the patient must be given comprehensive information about all stages.

Does fetal antigen 1 (FA1) identify cells with regenerative, endocrine and neuroendocrine potentials? A study of FA1 in embryonic, fetal, and placental tissue and in maternal circulation.

Fetal antigen 1 (FA1) is a circulating EGF multidomain glycoprotein. FA1 and its membrane-associated precursor is defined by the mRNAs referred to as delta-like (dlk), preadipocyte factor 1 (pref-1) or zona glomerulosa-specific factor (ZOG). Using a polyclonal antibody recognising both forms, the localisation of FA1/dlk was analysed in embryonic and fetal tissues between week 5 to 25 of gestation and related to germinal origin and development. FA1 was observed in endodermally derived hepatocytes, glandular cells of the pancreas anlage, and in respiratory epithelial cells. FA1 was also present in mesodermally derived cells of the renal proximal tubules, adrenal cortex, Leydig and Hilus cells of the testes and ovaries, fetal chondroblasts, and skeletal myotubes. Ectodermally derived neuro- and adenohypophysial cells, cells in the floor of the 3rd ventricle and plexus choroideus were also FA1 positive. The number of cells expressing FA1 decreased during fetal development where the expression became restricted to specific functional cells. Epidermis, gut epithelium, gall bladder, blood cells, spleen, thyroid gland, salivary glands, and smooth muscle cells were FA1 negative. Analysis of extra-embryonic tissues from normal and pathological pregnancies revealed FA1 in stromal cells surrounding the blood islands of the yolk sac as well as in placental fibroblasts where the expression was most pronounced in diploid, androgenic complete hydatidiform moles. However, as measured by ELISA, the circulating maternal FA1 levels in complete moles were not different from normal pregnancies. The results presented suggest that FA1 is a growth and/or differentiation factor extensively expressed in immature cells and down-regulated during fetal development. FA1 down-regulation was associated with a shift in the subcellular localisation indicating differential post-translational/post-transcriptional modifications during fetal development. FA1 may be a new marker of cellular subtypes with a regenerative potential and of specific cells with endocrine or neuroendocrine functions.

Complications and re-operation rate after tension-band wiring of olecranon fractures.

Tension-band wire fixation of olecranon fractures leads to a high re-operation rate because of the need to remove the metalware. This problem has commonly been thought to be related mainly to the backing-out of the Kirschner wires. A retrospective study was carried out in 55 patients with displaced olecranon fractures operated on with the tension-band wiring technique, in whom there was an overall 71.7% re-operation rate. Complications were few and minor in most patients. The main reason for the removal of the metalware was a direct complaint from the patient (in 61.3% of all removals). A literature review analyzing the causes of metalware removal is also presented.

Fetal antigen 1, an EGF multidomain protein in the sex hormone-producing cells of the gonads and the microenvironment of germ cells.

Fetal antigen 1 (FA1), an epidermal growth factor (EGF) multidomain glycoprotein, was investigated in the human reproductive system. Immunohistochemical analysis of the male reproductive system revealed staining for FA1 in the Leydig cells only. Concentrations of FA1 in seminal plasma and serum were similar and significantly correlated in weekly samples from three men (P < 0.0065). The concentrations in seminal plasma from vasectomized men (n = 4) were not significantly different from those of normal men (n = 187). The concentration of FA1 in seminal plasma was significantly correlated with the sperm counts of normozoospermic men (P < 0.0001), and significantly higher in seminal plasma from men with sperm counts > 20 x 10(6)/ml, compared with those with counts

Indirect measurement of mitochondrial proton leak and its application.

In mitochondria, ATP synthesis is coupled to oxygen consumption by the proton electrochemical gradient established across the mitochondrial inner membrane in a process termed oxidative phosphorylation. It has long been known from stoichiometric studies that ATP synthesis is not perfectly coupled to oxygen consumption. The major inefficiency in the system is leakage of protons across the mitochondrial inner membrane driven by the proton electrochemical gradient. The kinetics of the proton leak can be determined indirectly, by measuring the oxygen consumption of mitochondria under non-phosphorylating conditions (plus oligomycin) as a function of the proton electrochemical gradient. This experimental system provides a convenient means to investigate inner membrane permeability to protons and the effect of factors that may effect that permeability. In this paper we review some results from our laboratory of indirect measurement of mitochondrial proton leak and how it has been applied to investigate the effect of aging, obesity and thyroid status on proton leak. The results show that (i) proton leak in isolated liver mitochondria is not significantly different in a comparison of young and old rats, in contrast (ii) there is an apparent increase in proton leak in in situ mitochondria in hepatocytes from old rats when compared to those from young rats, (iii) proton leak in neuronal mitochondria in situ in synaptosomes is not significantly different in young and old rats, (iv) proton leak is greater in isolated liver mitochondria from ob/ob mice compared to lean controls, (v) acute leptin (OB protein) administration restores the increased leak rate in isolated liver mitochondria from ob/ob mice to that of lean controls, (vi) administration of thyroid hormone (T3) increases proton leak in rat muscle mitochondria, and (vii) proton leak in muscle mitochondria is insensitive to the presence of GDP. It is proposed that the experimental system described here for measuring proton leak, is an ideal functional assay for determining whether the novel uncoupling proteins increase inner membrane permeability to protons.

Fetal antigen 1, a member of the epidermal growth factor superfamily, in neurofibromas and serum from patients with neurofibromatosis type 1.

Fetal antigen 1 (FA1) is a 26-32 kDa glycoprotein containing six epidermal growth factor-like repeats closely related to the delta/notch/serrate proteins in Drosophila. FA1 has been shown to be involved in cell differentiation in a juxtacrine/paracrine manner. As neurofibromatosis type 1 (NF-1), also called von Recklinghausen disease, involves aberrant growth of tissues derived from the neural crest, the expression of FA1 was examined in neurofibroma skin biopsies and serum from patients with NF-1. FA1 was found in the spindle cells of all (n = 10) skin tumour specimens from adult NF-1 patients, whereas normal dermis was FA1 negative. In adults, the serum FA1 levels were significantly higher in NF-1 patients (n = 13) than in normal healthy controls (n = 177) (P = 0.037). In the group of children with NF-1 (n = 9), significantly higher serum FA1 levels were observed in those known to have complications with cerebral or spinal involvement (n = 4) (P = 0.014). The presence of FA1 in neurofibroma specimens and the elevated serum levels in patients with NF-1 suggests that FA1 may be involved in the pathogenesis of NF-1, perhaps acting as a growth promoting factor.

Thermal instability of the trimeric structure of the N-terminal propeptide of human procollagen type I in relation to assay technology.

The N-terminal propeptide of procollagen type I (PINP) appeared in two peaks after size chromatography. The high-molecular weight form was transformed to the low-molecular weight form during incubation at 37 degreesC, whereas the low-molecular weight form remained unchanged. The PINP concentrations in amniotic fluid and sera remained unchanged during 37 degreesC incubation when measured using an ELISA; however, concentrations decreased by 89-93% when measured using an RIA. The ELISA:RIA ratio varied from 1.1 to 2.9 in these fluids because of different size distributions and the inability of the RIA to measure the low-molecular weight form. Thermal transition of the high-molecular weight form caused a change in its elution volume but did not change its migration in sodium dodecyl sulfate-polyacrylamide gel electrophoresis. Mass spectrometry revealed identical results for both forms. We reached the following conclusions: (a) the trimeric structure of PINP is unstable at 37 degreesC; (b) the two molecular forms represent intact alpha1 chains in trimeric and monomeric forms; (c) thermal transition is an ongoing in vivo process; and (d) this is important in the choice of assay technology.

Knee injury and obesity in patients undergoing total knee replacement: a retrospective study in 115 patients.

The prevalence of obesity and previous knee injury was assessed in a retrospective study of 115 patients under-going total knee replacement due to osteoarthritis. Obesity was considered a contributing factor in the development of osteoarthritis in 37% of the patients, and 33% of the patients had had an injury to the knee in question. Unilateral osteoarthritis was significantly more frequent than bilateral osteoarthritis among patients with a history of previous knee injury. The association of previous injury to the knee and unilateral osteoarthritis was stronger in men than women. Aggressive treatment of patients with knee injuries seems warranted.

[Fatal poisoning with Letigen]. / Dødelig forgiftning med Letigen.

The anorexic agent Letigen, which contains 200 mg coffeine and 20 mg ephedrine, has been extensively used during the last decade. The case report describes a 19 year-old woman who ingested 50 Letigen tablets in a suicidal attempt. She developed severe toxic manifestations from the heart, CNS, muscles, liver and kidneys leading to several cardiac arrests, and died subsequently of cerebral oedema and incarceration on the fourth day of hospitalization. Because of the potentially life-threatening intoxication following an overdose, prescription of Letigen must be carefully administered.

Interlocking nailing of humeral shaft fractures.

: In this retrospective study 48 humeral shaft fractures in 48 patients were operated on using the Seidel interlocking nail. The length of follow-up ranged from 6 to 60 months (median, 26 months). The treatment of fractures was satisfactory with the Seidel nail, but we emphasize the importance of countersinking the tip of the nail into the humeral head to avoid impingement. In 5 of 12 patients with non-unions, the procedure failed, and we found that the distal locking seemed to be inadequate. Pathological fractures (i.e., those caused by metastatic tumors) were all efficiently treated with the Seidel nail.