The healthy donor effect impacts self-reported physical and mental health - results from the Danish Blood Donor Study (DBDS).

AIMS: This study aimed at quantifying the healthy donor effect by comparing self-perceived mental and physical health between blood donors and non-donors. BACKGROUND: In theory, the selection process known as the healthy donor effect should result in better self-perceived, health-related quality of life in donors than in non-donors. METHODS: The Short Form-12 data from the Danish Twin Registry (DTR) was compared with the data from the Danish Blood Donor Study (DBDS). Data on age, sex and smoking status were included in the analyses. The multivariable linear regression analysis was stratified by sex and age group intervals. Outcome variables were the mental component score (MCS) and the physical component score (PCS). RESULTS: A total of 28 982 and 36 913 participants from the DTR and the DBDS, respectively, were included in this study. Younger donors had higher MCS than non-donors, whereas MCS was only marginally high in older donors compared with non-donors. In contrast, PCS was almost similar for both young donors and non-donors. With the increase in age, non-donors had lower PCS than donors. CONCLUSIONS: Two selection patterns were revealed. Among young individuals, better self-perceived mental health was associated with a blood donor. With the increase in age, better self-perceived physical health was associated with blood donation.

Long-term effects of smallpox vaccination on expression of the HIV-1 co-receptor CCR5 in women.

BACKGROUND: Smallpox vaccinations were stopped globally in 1980. Recent studies have shown that in women, being smallpox vaccinated was associated with a reduced risk of HIV infection compared with not being smallpox vaccinated. At the initial infection, HIV-1 most often uses CCR5 as a co-receptor to infect the T-lymphocytes. We therefore investigated whether smallpox vaccination is associated with a down-regulation of CCR5 on the surface of peripheral T-lymphocytes in healthy women in Guinea-Bissau. METHODS: We included HIV seronegative women from Bissau, Guinea-Bissau, born before 1974, with and without a smallpox vaccination scar. Blood samples were stabilised in a TransFix buffer solution and stained for flow cytometry according to a T-cell maturation profile. RESULTS: Ninety-seven women were included in the study; 52 with a smallpox vaccination scar and 45 without a scar. No association between smallpox vaccination scar and CCR5 expression was found in any T-lymphocyte subtype. CONCLUSION: Among HIV seronegative women, being smallpox vaccinated more than 40 years ago was not associated with a down-regulation of CCR5 receptors on the surface of peripheral T-lymphocytes.

Low-grade inflammation is associated with lower haemoglobin levels in healthy individuals: results from the Danish blood donor study.

BACKGROUND AND OBJECTIVES: Chronic inflammation can lead to anaemia of chronic disease due to the sequestration of iron caused by inflammatory cytokines and the protein hepcidin. However, the effect of low-grade inflammation (LGI) on haemoglobin among healthy individuals is not known. This study examines the effect of LGI on haemoglobin among Danish blood donors. MATERIALS AND METHODS: We performed multivariable linear regression to assess the effect of LGI (i.e. high-sensitivity C-reactive protein above 3 mg/l but below 10 mg/l) on haemoglobin in 17 322 Danish blood donors. We also performed multivariable logistic regression to evaluate the effect of LGI on the risk of having low haemoglobin (below the 10th percentile among men and women, respectively). We adjusted for donation activity, age, sex, low ferritin, oral contraceptives and menopause. All analyses were stratified by current smoking status. RESULTS: LGI was associated with lower haemoglobin (0·08 mm lower [0·12 g/dl], 95% confidence interval (CI): -0·11-0·05) and increased risk of low haemoglobin (OR = 1·22, 95% CI: 1·05-1·43) in non-smokers. Conversely, LGI was associated with higher haemoglobin in smokers (0·12 mm [0·19 g/dl], 95% CI: 0·06-0·18). CONCLUSION: In this first study of LGI and haemoglobin in healthy individuals, there was a negative association between LGI and haemoglobin in non-smokers. The association was positive in smokers, probably because smoking leads to both increased inflammation and increased haemoglobin through CO exposure.

Spermatogenic capacity in fertile men with elevated exposure to polychlorinated biphenyls.

BACKGROUND: Endocrine disrupting industrial chemicals, such as polychlorinated biphenyls (PCBs), are suspected to adversely affect male reproductive functions. OBJECTIVES: The Faroe Islands community exhibits an unusually wide range of exposures to dietary contaminants, and in this setting we examined the possible association between PCB exposure and semen quality and reproductive hormones in fertile Faroese men. METHODS: Participants in this cross-sectional study include 266 proven fertile men residing in the Faroe Islands. PCB levels and hormone profiles were measured in serum samples taken at the clinical examination that included semen quality parameters. RESULTS: A significant positive association was seen between serum-PCB and the testosterone/estradiol ratio (p=0.04). In the unadjusted analyses, elevated PCB exposure was associated with increased serum concentrations of SHBG (p=0.01) and FSH (p=0.05). We found no association between the serum PCB concentration and the semen quality variables. CONCLUSION: In this population of highly exposed fertile men, the current serum-PCB concentration was associated with higher androgen/estrogen ratio. Further studies are needed to establish the findings and further document PCB-associated hormonal effects, any time windows of increased susceptibility, and the role of PCB in sub-fecundity.

Case report: binding of a clinically relevant human leukocyte antigen-DQα-specific antibody in a kidney graft recipient is inhibited by donor-type human leukocyte antigen-DQß chain.

In this case report, we have found what may be an immunization with donor-specific human leukocyte antigen (HLA)-DQα in combination with recipient-specific HLA-DQß. A renal allograft recipient who did not comply with immunosuppressive therapy during pregnancy had graft failure 23 months posttransplantation with biopsy-proven humoral and cellular rejection. Sera were tested in a Luminex-based single-antigen bead assay. We compared Luminex reactivity with the degree of eplet mismatching between the recipient's own HLA-DQ chains and the HLA-DQ chains bound to the Luminex beads. Eplet calculations were done with the HLAMatchmaker. HLA-DQ similarities were compared further by dissimilarity scoring in HistoCheck. We observed that Luminex beads with donor-type HLA-DQα and HLA-DQß bound less antibody than beads with donor-type HLA-DQα combined with recipient HLA-DQß. In HLAMatchmaker, we identified all eplet mismatches between donor and recipient HLA-DQ. Next, we counted how many of these eplets were represented on the various Luminex beads. We found that antibody binding to the bead increased with the number of such mismatches for HLA-DQα. Surprisingly, antibody binding decreased as the number of eplet mismatches for HLA-DQß increased, from a mean fluorescence intensity (MFI) value of 18,800 for no mismatched eplets to approximately 10,000 for 12 mismatched eplets. These findings were confirmed by comparing antibody binding with the structural dissimilarity score between the recipient HLA-DQ type and the HLA-DQ bound to the Luminex beads. In this patient, clinically relevant antibodies bound strongly to donor-like HLA-DQα chains when combined with recipient-like HLA-DQß. HLA-DQß chains more similar to those of the donor reduced the binding of donor- specific HLA-DQα antibody.

Effect of remote ischemic conditioning on dendritic cell number in blood after renal transplantation--flow cytometry in a porcine model.

Delayed graft function after transplantation increases the risk of rejection. Remote ischemic conditioning (rIC) consists of repetitive, brief, non-damaging periods of ischemia in a limb. For reasons not fully understood, rIC protects the target organ against subsequent ischemia-reperfusion injury. Because ischemic endothelium attracts dendritic cells (DCs), we hypothesised that rIC protects the organ by "trapping" circulating DCs in the limb exposed to rIC. With fewer DCs thus available to infiltrate the graft, a strong T-cell mediated immune response toward the graft is less likely. To test this hypothesis, we measured the number of circulating DCs in a porcine model of renal transplantation with and without rIC. Brain death was induced in eight 65-kg donor pigs. After 22 h of cold ischemia, the kidneys were transplanted into sixteen 15-kg recipient pigs. The recipients were randomised to either non-rIC or rIC before reperfusion of the graft and observed 10 h after reperfusion. The number of DCs was determined by flow cytometry. DCs were identified on the basis of forward- and side-scatter characteristics of CD14-negative mononuclear cells with expression of CD172a. Dendritic cells were subclassified as either plasmacytoid (pDCs) (CD172a(dim), CD4(+), CD14(-)) or conventional (cDCs) (CD172a(high), CD4(-), CD14(-)). Remote ischemic conditioning did not affect the number of circulating cDCs or pDCs within the 10h after transplantation studied. Regardless of rIC, the number of pDCs decreased after graft reperfusion and then returned to baseline levels. In contrast, the number of circulating cDCs increased after reperfusion and later returned to baseline levels.

A simple method for unbiased quantitation of adoptively transferred cells in solid tissues.

In a mouse model, we demonstrate how to obtain a direct, unbiased estimate of the total number of adoptively transferred cells in a variety of organs at different time points. The estimate is obtained by a straightforward method based on the optical fractionator principle. Specifically, non-stimulated C57BL/6J mouse splenocytes were labelled with carboxyfluorescein diacetate succinimidyl ester (CFSE) and adoptively transferred to normal C57BL/6J mice by intravenous injection. The total number of CFSE-positive cells was subsequently determined in lung, spleen, liver, kidney, and inguinal lymph node at six different time points following adoptive transfer (from 60 s to 1 week), providing a quantitative estimate of the organ distribution of the transferred cells over time. These estimates were obtained by microscopy of uniform samples of thick sections from the respective organs. Importantly, the samples were chosen and prepared in accordance with the optical fractionator principle. We demonstrate that the method is simple, precise, and well suited for quantitative immunological studies.

Effect of dendritic cells on flow cytometric quantification of cytomegalovirus-specific, interferon-gamma-producing T cells.

Flow cytometric measurement of intracellular cytokines in T cells exposed to antigen is a widely used method for quantification of an antigen-specific T-cell response. As the frequency of antigen-specific T cells is often very low, any improvement in signal to noise ratio is of great importance. Thus, in this study, the ability of antigen-pulsed dendritic cells (DCs) to increase the number of antigen-specific, interferon-gamma (IFN-gamma)-producing CD4+ T cells measurable both in fresh peripheral blood and in reconstituted frozen blood mononuclear cell (MNC) samples was evaluated. Cytomegalovirus (CMV) was used as antigen in a 10 h assay, using cells from both CMV-seropositive and -seronegative donors. When reconstituted frozen samples were analysed, the general response towards CMV lysate in CMV-seropositive donors was 23-86% lower compared to the corresponding fresh blood samples. Antigen-pulsed DCs could not improve the sensitivity of the intracellular cytokine-detection assay when fresh peripheral blood samples were used. Interestingly, however, the addition of CMV lysate-pulsed DCs to cryopreserved MNC samples substantially increased the frequency of specifically induced IFN-gamma-producing cells to a level comparable to the frequency found in the corresponding fresh blood samples.

Influence of surgery and endotoxin-induced sepsis combined on natural killer cell activity, oxidative burst of granulocytes and antigen presentation capability of monocytes.

BACKGROUND: Cell mediated immunity is affected in the course of sepsis and following surgical stress. The natural killer (NK) cells, the granulocytes and the monocytes constitute the immediate unspecific cell mediated immunity. We therefore investigated the effect of surgery- and endotoxin-induced sepsis on NK cells, granulocytes and monocytes in a two-hit model. METHODS: Three groups of 40 mice. Each group was divided into four groups of 10 mice. All the animals were anesthetized and subjected to either: laparotomy; treatment with Escherichia coli endotoxin i.p.; laparotomy followed 20 min later by endotoxin i.p.; or left untreated as a control group. In the first 40 mice the NK cell activity in the spleen and number of NK cells in the liver were measured, in the second the oxidative burst of granulocytes, and in the third the antigen presentation capacity of monocytes. RESULTS: Endotoxin stimulated the NK cell activity and up-regulated the antigen presentation capability on monocytes. In contrast, surgical stress reduced the NK cell activity, the number of NK cells and down-regulated the antigen presentation capability on monocytes. After surgery, followed by administration of endotoxin, the oxidative burst of granulocytes was stimulated while antigen presentation capability on monocytes was down-regulated. Endotoxin prevented or reverted the postoperative suppression of NK cell activity. CONCLUSION: Our two-hit model shows that some cell types of the unspecific immune system exhibit an excessive inflammatory response (NK cells, granulocytes) while specific functions of other cell types (monocytes) are simultaneously diminished. This diversity makes a potential therapeutic immunomodulation very complex as some cell types would need to be down-regulated while others need to be stimulated.

Transgene expression in human epidermal keratinocytes: cell cycle arrest of productively transfected cells.

We have analysed the consequences of liposome mediated gene transfer into human primary epidermal keratinocytes and compared non-Epstein-Barr Virus (EBV) and EBV based expression vectors that carry the genes encoding human Growth Hormone (hGH) or Enhanced Green Fluorescent Protein (EGFP). Different kinetics between the non-EBV and EBV based vectors were revealed upon subcultivation of hGH transfected keratinocytes. The keratinocytes transfected with non-EBV based vector showed a rapid reduction in hGH production. Although the EBV based vector resulted in more stable expression, this was also reduced over time. Chromatin inactivation by deacetylation was investigated by treatment with sodium butyrate and found not to be the reason for the decreasing expression. Keratinocytes divided into subpopulations enriched for either stem cells or transit amplifying cells, based on beta1-integrin expression and function, do not differ significantly with respect to susceptibility to productive transfection. However, when the keratinocytes were transfected with the EGFP gene and sorted live by FACS into EGFP negative and positive populations, only the negative cells were capable of forming significant numbers of colonies. This is consistent with the observation that the ability to incorporate BrdU was dramatically reduced in the EGFP expressing population within 24-48 h post transfection indicating an almost complete cell cycle arrest. p53 levels were unaffected by the procedures, and the keratinocyte cell line HaCat, mutated in both p53 alleles, also shows a marked reduction in clonogenic potency upon transfection. There was a slight increase of TUNEL positive apoptotic nuclei in the positive population at early time points. However, the apoptotic index was still very low. When we measured the frequency of involucrin expressing cells, we found an increase in the productively transfected population over time indicating an initiation of terminal differentiation. In contrast to the transfected cultures, keratinocytes that were transduced using a retroviral vector showed no decrease in colony forming efficiency. In conclusion we find that transgene expressing cells from transfected cultures of epidermal keratinocytes undergo cell cycle arrest and initiate terminal differentiation by mechanisms which are independent of p53 levels.

Characterization of murine dendritic cells derived from adherent blood mononuclear cells in vitro.

The therapeutic potential of dendritic cells loaded with tumour antigens for the induction of effective immune responses against cancer is currently being tested in numerous clinical trials. In most cases, the dendritic cells are generated in vitro from peripheral blood monocytes. Many aspects of dendritic cell-based vaccination have not yet been examined in detail, and homologous mouse model systems may prove very valuable for optimizing clinical procedures. In the murine system, however, dendritic cells are usually isolated from either lymphoid tissues or bone marrow cultures. To date, murine monocyte-derived dendritic cells have been described only sporadically. Here, we describe a culture system for the generation of murine dendritic cells from adherent peripheral blood mononuclear cells by culturing in the presence of granulocyte-macrophage colony stimulating factor and interleukin-4. After 7 days of culture the nonadherent cells were harvested from the cultures. Most of these cells exhibited well-accepted characteristics of mature dendritic cells (e.g. veiled appearance, high expression of major histocompatibility complex class II and CD86) and stimulated vigorous proliferation of allogeneic T cells in a primary mixed leucocyte reaction following stimulation with bacterial lipopolysaccharide. Interestingly, staining the cells for expression of the putative antigen-uptake receptor DEC-205 revealed a distinct bimodal distribution.

Strain-specific variations in the development of dendritic cells in murine bone-marrow cultures.

The dendritic cell (DC) is a professional antigen-presenting cell of central importance in immunity. In this paper, we examined DCs generated by 11-day culture of bone-marrow cells from the four mouse strains C57BL/6J, BALB/cA, C3H/HeN and B10.PL-H2u (73NS)/Sn with respect to cell yield as well as surface-marker phenotype and morphology. We also investigated the phenotypic changes and the T-cell stimulatory activity of the DCs induced by bacterial lipopolysaccharide (LPS). Morphologically, we observed low levels (5-10%) of granulocyte contamination of the cultures after a culture period of 11 days. Considerable strain-specific differences were found in the expression levels of the surface markers in addition to the differences in the ratio of the immature to mature DCs in the cultures that were not stimulated with LPS. Furthermore, we found that LPS strongly induces maturation of DCs in all strains investigated with the exception of the B10.PL strain.

Treatment of hyperextension injuries to the PIP joint.

A prospective randomized trial of type 1 hyperextension injuries to the PIP joint treated conservatively by an elastic double-finger bandage or an aluminium splint for 2 weeks showed no differences in the clinical outcome after 6 months.

Biomechanical evaluation of distal radioulnar reconstructions.

Numerous reconstructive procedures have been described for the treatment of chronic instability of the distal radioulnar joint or instability of the stump of the resected distal ulna. This biomechanical study presents an evaluation of the three basic design types that have been used in reconstruction. The initial static stability provided by the reconstructions was tested and compared with the stability of the intact joint. Our findings show that all reconstructive procedures failed to restore natural joint stability. A radioulnar sling design was the most effective of the three types, whereas tenodesis procedures and ulnar collateral ligament reconstruction were much less effective in providing stability. We conclude that current designs have significant biomechanical shortcomings. On the basis of our observations during testing, we believe that improved designs will require an intra-articular reconstruction that more closely duplicates the biomechanical functions of the triangular fibrocartilage complex.

Postoperative urinary retention associated with total hip and total knee arthroplasties.

Urinary retention following total hip and knee arthroplasty is a common problem frequently necessitating either prolonged urethral catheter drainage or intermittent catheterization. The direct relationship of urinary tract instrumentations, procedures, and infections to deep sepsis in total hip replacements is well documented. Pharmacologic therapy to stimulate voiding or augment bladder emptying is thus theoretically preferable to the use of catheterization. Prasozin hydrochloride, an alpha blocker, relaxes the smooth musculature of the posterior urethra and prostrate and has been used to treat urinary obstruction secondary to benign prostatic hypertrophy. A prospective study in 60 male patients showed a statistically significant decrease in postoperative urinary retention with the perioperative administration of prazosin (p less than 0.01). A higher incidence of urinary tract infection was seen in patients who developed urinary retention (3% versus 20%) (p less than 0.01). Uroflowmetry parameters were not predictive of the patient at risk for retention and were unaltered with the administration of prazosin. Prazosin can be an effective adjunct in the prophylaxis of postoperative urinary tract infections and may decrease the potential risk for total joint sepsis.