Rectum perforation during transanal irrigation: a case story.

STUDY DESIGN: Case report. OBJECTIVES: Report a case of rectum perforation during transanal irrigation (TAI). SETTING: Clinic for Spinal Cord Injuries, and Departments of Gastroenterological Surgery and Radiology. CASE REPORT: A 54-year-old woman with spinal cord lesion for 35 years emptied for years her bowel using oral laxative. This became more difficult and took more than 2 h three times a week with reflex stimulation after a chlysma. She wanted to try TAI, and went through the procedure with a nurse one time. The next time she performed the TAI by herself without difficulty. Two hours later she started shivering with a temperature at 38.3 degrees C with normal blood pressure (BP). At rectal exploration, a spoonful fresh blood was found. After another 2 h, she became septic and was transferred to a gastroenterological surgical department. An abdominal X-ray gave no suspicion of free air in the abdomen. Sigmoideoscopy showed 3-5 cm oral to the dentate line a 1 x 1 cm transmural circular lesion. A colon X-ray with water contrast showed a perforation of approximately 2 x 0.5 cm. CONCLUSION: Even in experienced individuals who are proper trained, TAI can cause rectal perforation, which always have to be born in mind.

Haematopoietic cell transplantation with non-myeloablative conditioning in Denmark: disease-specific outcome, complications and hospitalization requirements of the first 100 transplants.

We analysed the outcome and hospitalization requirements of the first 100 patients (Hodgkin's disease (HD), N=13; multiple myeloma (MM), N=14; CLL, N=12; non-Hodgkin's lymphoma (NHL), N=17; myelodysplastic syndrome (MDS), N=18; AML, N=24 and CML, N=2) treated in Denmark with haematopoietic cell transplantation after non-myeloablative conditioning with TBI 2 Gy+/-fludarabine. The cumulative incidence of acute GVHD grade II-IV and extensive chronic GVHD was 67 and 49%. After a median follow-up of 534 days, the overall survival, PFS, relapse-related mortality and treatment-related mortality were 59, 50, 25 and 17%, respectively. Patients with CLL, NHL, AML and MDS with <5% blasts at any time had a favourable outcome with a PFS of 61-71%. Patients with MM, HD and MDS and a history of > or =5% blasts had a less favourable outcome with a PFS of 19-38% (P=0.001). The cumulative incidence of discontinuation of immunosuppression was 37%. During the first and second year post transplant, patients experienced a mean of 41 and 13 outpatient clinic visits, and 53 and 16 days of hospitalization. Sixteen patients were admitted to the intensive care unit, of whom eight are still alive. In conclusion, transplantation outcomes were encouraging, but complications requiring admission and outpatient clinic visits occur frequently post transplant.

Stromal-mediated down-regulation of CD13 in bone marrow cells originating from acute myeloid leukemia patients.

The metallopeptidase CD13 is expressed on normal myeloid cells of monocytic and granulocytic origin and on the surface of leukemic blasts in most acute myeloid leukemias (AML). To study the mechanisms regulating lineage restricted CD13 expression in AML we determined normalised CD13 mRNA levels in bone marrow cells and peripheral blood cells of 27 AML patients. Cells of bone marrow origin had lower levels of normalised CD13 mRNA than cells of peripheral blood origin, even though fluorescence intensity and fraction of cells expressing CD13 on the surface was unchanged. In particular, AML patients with very low levels of normalised CD13 mRNA in bone marrow cells showed an increase in CD13 mRNA expression in peripheral blood. To evaluate the effects of bone marrow microenvironment on CD13 mRNA expression, we cultured leukemic myeloid cells with and without murine stromal cells. Bone marrow cells with high and low CD13 surface expression that entered the stromal layers all down-regulated CD13 mRNA expression as compared to cells in suspension above. For peripheral blood cells within stromal layers, CD13 mRNA expression was diminished in only 3 out of 6 cases. The ambiguous effect of stromal cells on peripheral blood cells may illustrate a differentiation-dependent response towards stroma. We determined the polyadenylation status of CD13 mRNA for 9 bone marrow aspirates and 7 peripheral blood samples. Polyadenylation was diminished in bone marrow cells from AML patients with low levels of normalised CD13 mRNA, raising the possibility of involvement of mRNA instability in regulation of CD13 mRNA expression in this subgroup of patients.

FAB M4 and high CD14 surface expression is associated with high cellular resistance to Ara-C and daunorubicin: implications for clinical outcome in acute myeloid leukaemia.

In 145 adult patients diagnosed with non-M3 acute myeloid leukaemia (AML) the relevance of FAB-subtype and immunophenotype to in vitro cellular drug resistance towards the anthracyclines aclarubicin (Acla) and daunorubicin (Dau), and the nucleoside analogue cytarabine (Ara-C), as well as other antileukaemic drugs, was investigated using a 4-d MTT (3-[4,5-dimethylthiazol-2-yl]-2,5-diphenyl tetrazolium bromide) assay. We demonstrate that high CD14 expression is highly significantly associated with high cellular Ara-C and Dau resistance in univariate as well as multivariate analyses. FAB subtypes with highest and lowest cellular Ara-C resistance were M4 and M5, respectively (P < 0.01, one-way anova), whereas FAB subtypes with highest and lowest cellular Dau resistance were M4 and M1, respectively (P < 0.01, one-way anova). By contrast, no significant differences in cellular drug resistance towards Acla could be demonstrated among FAB subtypes. Furthermore, in two cohorts of AML patients treated by two different regimens for remission induction over a period of 15 yr (1985-94, n = 159 and 1995-99, n = 76, respectively) we demonstrate in univariate analyses a significance of CD14 expression with respect to clinical outcome. With the exception of significance to probability of obtaining complete remission in the first cohort (P = 0.03, logistic regression), this significance was, however, lost in multivariate analyses. It was demonstrated that FAB-M4 patients were older than M5 patients and that high CD14 expression was associated with the presence of secondary AML and older age. We conclude that although cases with high blast cell CD14 expression (and FAB-M4 cases) were more resistant to Ara-C as well as Dau in vitro, the clinical and biological significance of this may be debatable because of interactions with major prognostic factors in AML.

Long-term culture of hematopoietic stem cells - validating the stromal component of the CAFC assay.

BACKGROUND: The stroma-based long-term culture is the assay of choice when a functional detection of primitive hematopoietic cells in vitro is sought. However, different stromal cell lines varying in supporting capacity have been raised and applied in different laboratories, resulting in a wide range in published frequencies of LTCIC alternative CAFC. METHODS: In order to identify the most suitable stromal source in terms of supportive capacity, reproducibility, and ease of handling, we have compared some of the most commonly employed murine cell lines to human bone marrow stroma in secondary long-term culture set-ups. RESULTS: Seeking an approximation to the supportive capacity of human BM stroma we found the FBMD-1 cell line supplemented with G-CSF and IL-3 superior to FBMD-1 cells alone, and to M2-10B4 and Sl/Sl cells. Moreover, in co-cultures of CD34(+) cells and the FBMD-1 line, we found week 5 CAFC content highly reproducible (50.5 +/- 6.66 - 54.6 +/- 7.07/10(4) plated cells, p value > 0.95) and the assay was suitable for inter-individual comparison in a clinical setting. In fact, the week 5 CAFC results were even more reproducible than those of the CFU assays (CV 0.03 for the CAFC assay versus 0.13-0.33 for the CFU assays). On the other hand, when extending the culture period to 8 weeks, the cobblestone area formation was best maintained by human BM stroma and the high reproducibility in CAFC enumeration in cultures supported by the FBMD-1 was lost. DISCUSSION: Among the stromal cell sources tested, the FBMD-1 line was found to be superior in terms of ease of handling and week 5 CAFC reproducibility. However, this robustness could not be extended to week 8 CAFC.

[Hematopoietic stem cells I. Basal aspects]. / Haematopoietiske stamceller I. Basale aspekter.

The knowledge on basic aspects of hematopoietic stem cells (HSC) has markedly increased during the last decade to the extent that such cells are now routinely employed therapeutically in order to restore hematopoiesis following myeloablative chemotherapy and irradiation in patients with malignant disorders. Different methods can be applied to characterize HSC. Thus, by immunophenotyping and flow cytometry it is possible to delineate subpopulations of cells enriched for HSC. However, since phenotypic characteristics associated with HSC do not necessarily correlate to their developmental potential, measurements of HSC also rely heavily upon functional in vitro and in vivo assays, which demonstrate the presence of HSC through their ability to proliferate and differentiate in a clonogenic fashion. In this review we describe the assays developed to characterize and quantitate immature HSC, which form the basis for their clinical application.

[Hematopoietic stem cells II. Diagnostic and therapeutic aspects]. / Haematopoietiske stamceller II. Diagnostiske og terapeutiske aspekter.

Characterization and isolation of haematopoietic stem cells (HSC) have resulted in their clinical application in patients with malignant disorders and--through gene therapeutic initiatives--also in the treatment of inherited diseases. Autologous stem cell transplantation (ASCT), which was introduced because of the high number of relapses in cancer patients in remission, involves dose-intensification (conditioning), which induces myeloablation. In this setting, reinfusion of HSC is performed to restore haematopoiesis. Flow cytometric determination of CD34+ cells and clonogenic assays for committed myeloid HSC (CFU-GM) are vehicles for quality control of the harvested HSC material and are integrated into the ASCT programs. Moreover, harvest of HSC and purification of CD34+ cells enables new treatment options such as removal of cancer cells from grafts, optimization of gene transduction as well as ex vivo expansion of HSC before reinfusion. In conclusion, the expanding insights into HSC in the 1990's have already been translated into valuable diagnostic and therapeutic modalities.

Danish patients with untreated multiple myeloma do not harbour human herpesvirus 8.

The role of human herpesvirus 8 (HHV-8) in multiple myeloma (MM) remains controversial. We examined 15 Danish MM patients before cytoreductive therapy. Mononuclear cells isolated from peripheral blood and bone marrow aspirates, as well as long-term cultured bone marrow stromal cells, were assayed for the presence of HHV-8 DNA. All material was tested by three simple unnested polymerase chain reaction (PCR) assays (amplifying regions of ORF26, ORFK1 and ORF75) and two nested PCR assays (amplifying regions of ORF26). HHV-8 was not demonstrated in any of the samples. Our findings do not suggest an association between HHV-8 and MM in the Danish population.

Reduced total number of cobblestone area forming cells and in vitro stromal-cell growth in autografts from acute myeloid leukemia patients.

BACKGROUND: It is well known that ABMT in acute myeloid leukemia (AML) often results in delayed hematopoietic engraftment, but the reason behind this has not been resolved. Previous studies have largely dealt with measurements of committed myeloid progenitors as surrogate markers for hematopoiesis. METHODS: Measurements of Week 5 cobblestone area forming cells (CAFC) and stromal-cell growth in BM autografts from 14 AML patients were compared with those from 10 NHL patients. RESULTS: Grafts achieved from the AML patients contained a significantly lower total number of CAFC than those from the NHL patients. The reason for this was a lower total amount of mononuclear cells (MNC) obtained during harvest procedure (mean 0.4 x 10(8)/kg for AML, versus 0.8 x 10(8)/kg for NHL). In contrast, the frequency of CAFC was comparable both between patient groups (mean 1.47, range 0.15-6.33 per 10(4) MNC for AML versus mean 1.47, range 0.53-3.57 per 10(4) MNC for NHL) and compared with that of eight normal donors (mean 1.12, range 0.73-1.73 per 10(4) MNC). An inverse relationship was observed between the total CAFC number in the grafts and the hematopoietic reconstitution of both granulocytes > or = 2.0 x 10(9)/L and thrombocytes > or = 50 x 10(9)/L, in which the level of 9.0 x 10(3) CAFC/kg implied a prompt engraftment for both patient groups. Whereas the stromal cell outgrowth in vitro from 8/10 NHL patients was similar to that of six normal donors, only a few stromal cells appeared in the majority of nine evaluable AML patients. DISCUSSION: A decreased total CAFC content, as well as an inferior stromal-cell function, may be critical elements for prolonged hematopoietic reconstitution in AML.

Leukaemia cell drug resistance and prognostic factors in AML.

In 93 cases of acute myeloid leukaemia (AML) the extent to which prognostic factors mirrored the in vitro cellular chemotherapy resistance (to anthracyclines aclarubicin (Acla) and daunorubicin (Dau) as well as nucleoside analogue cytarabine (Ara-C)) was investigated using a 4-d MTT (3-[4,5-dimethylthiazol-2-yl]-2,5-diphenyl tetrazolium bromide) assay. We found that age at presentation and presence of secondary AML were significantly correlated to leukaemia cell Ara-C resistance. Thus, analysis of in vitro drug resistance data revealed that age at presentation and presence of secondary leukaemia were both independently correlated to cellular drug resistance, with older age being associated with higher Ara-C resistance in vitro (p=0.02 and 0.01 in univariate and multivariate analyses, respectively) and with secondary leukaemia being associated with higher Ara-C resistance (p=0.04 and 0.059 in univariate and multivariate analysis, respectively). Median LC-50 values (Ara-C) were: 178 ng/ml in paediatric cases, 356 ng/ml in younger adult cases, and 584 ng/ml in elderly (age > or = 60 yr) cases giving a resistance ratio between these age subgroups of 1:2.0:3.3. Median LC-50 values (Ara-C) was 381 ng/ml in de novo cases as opposed to 891 ng/ml (resistance ratio 1:2.3) in secondary cases. By contrast, cytogenetic findings, presenting leucocyte count, FAB-subtype, and gender were not consistently correlated to in vitro drug resistance to any of the three drugs. We conclude that at least two major adverse prognostic factors in AML (advanced age at presentation and presence of secondary leukaemia) are associated with increased leukaemia cell Ara-C resistance. High leucocyte count is not associated with increased cellular drug resistance towards Acla, Ara-C or Dau.

[Acute myeloid leukemia. Therapeutic results 1985-1994]. / Akut myeloid leukaemi. Behandlingsresultater 1985-1994.

In a retrospective study we evaluate the treatment and outcome of 421 adults admitted to our department with acute myeloid leukemia (AML) during the 10 year period from 1985-1994. Younger patients (< or = 55 years) had a significantly better prognosis than elderly patients (> 55 years), partly because more younger patients had remission-induction therapy (81% versus 39%) and their complete remission (CR) rate was higher (69% versus 41%). In patients achieving CR the long-term survival (five years) was 40% for younger and 26% for elderly patients. Nineteen patients received autologous bone marrow transplantation (BMT) and eight an allogeneic BMT in first CR. Five patients treated with allogeneic BMT are still relapse-free, whereas autologous BMT, in comparison to conventional chemotherapy alone, did not improve the five-year relapse-free survival (29 versus 27%). Our data illustrate that in a non-selected material of AML patients the long-term survival (five years) has only slightly improved. The effect of autologous BMT cannot be evaluated in this retrospective setting.

Delineation of erythropoiesis in normal and malignant bone marrow using monoclonal antibody AS-E1 directed against transferrin receptors (CD71).

We have delineated the erythropoietic compartment in normal and malignant bone marrow (BM) by using the monoclonal antibody (mAb) AS-E1 directed against the transferrin receptor by flow cytometric (FCM) analysis. In normal BM we found a bimodal expression in antigen density with a minor subset (approximately 3%) expressing AS-E1high and a larger subset (approximately 15%) expressing AS-E1low. By fluorescence activated cell sorting, morphological examination of smears stained by immunocytochemistry and by BFU-E assays the AS-E1high fraction was shown to contain cells of erythroid origin (proerythroblasts, basophilic erythroblasts and polychromatic erythroblasts), whereas the AS-E1low fraction consisted mainly of promyelocytes and myelocytes. In patients with malignant hematological disorders we found a more pronounced heterogeneity in the density and the degree of AS-E1low expression compared to normal BM, and to further characterize the AS-E1low cells in patients and to exclude that this broad reactivity interfered with the identification of the AS-E1high cells, we employed triple-color FCM assays with mAbs directed against the myeloid surface markers CD13 and CD66 in addition to AS-E1. In all patients we found that 80-90% of the AS-E1low cells co-expressed CD13 and/or CD66 and thus were of myeloid origin. Finally, we evaluated 2 methods for determination of the AS-E1high subset and found an assay involving forward light scatter and logAS-E1 density to be sufficient. We conclude that AS-E1high is a valid FCM marker for the normal erythropoiesis.

[Preventive strategies against infections in autologous bone marrow transplantation]. / Infektionsstrategi ved autolog knoglemarvstransplantation.

The routine microbial surveillance, the prophylatic antibiotic regime and the management of infections were retrospectively evaluated in 26 autologous bone marrow transplantation episodes. The surveillance specimens indicated that ciprofloxacin prophylaxis was effective in minimising gram-negative functions. The comparison between specimens taken before and during the granulocytopenic period showed a shift towards more yeast, otherwise there were no big differences. The surveillance specimen could not identify the causative organisms in case of fever. The proven infections comprised four cases of bacteriaemia and one case of gastro-enteritis. Two patients died from disseminated Aspergillus infection not diagnosed prior to death. In 11 episodes the cause of fever remained unknown. Both fungal infections were related to nearby building construction work. None of the patients with bacterial infections were in a serious clinical condition. In the future the number of routine microbial surveillance specimens will be reduced. The aim of surveillance will be to monitor the potential occurrence of fluoroquinolone resistance, and more attention will be paid to possible fungal infections. The antibacterial policy will be continued.

Chronic neutrophil leukaemia in adolescence and young adulthood.

Chronic neutrophil leukaemia (CNL) is a rare myeloproliferative disorder predominantly reported in elderly patients. We present a 15-year-old girl and a 25-year-old male with CNL. Clonal cytogenetic abnormalities were detected in both patients. One showed trisomy 21 evolving into tetrasomy 21. The second patient showed a unique chromosome aberration during blast crisis: t(2;2)(q32;p24). Both patients were successfully treated with allogeneic bone marrow transplantation (BMT). CNL should also be considered as a differential diagnosis in adolescence and young adulthood. BMT represents a potentially curative treatment option in such patients.

Acute admissions to medical departments. A comparison between an urban and a rural district.

To compare hospitalization into medical departments, acute admissions into a city hospital and into a district hospital were compared prospectively over a two-week period. Patients referred to the city hospital were on average older, were more frequently living alone and they had a greater amount of social care attendance in their homes. On the other hand, distribution of referral diagnoses, overall patient activity, occupational status and contact with relatives were similar in the two areas. Sub-acute or acute illness was considered the main cause of admission in both areas; the amount of admissions for social reasons was 13 percent to the city hospital versus 3 percent to the district hospital. Relevant alternatives to hospitalization seemed to exist in 50 percent of the admissions to the city hospital versus only 3 percent to the district hospital. Since patients admitted for social reasons block hospital beds for a longer time period than those admitted for other reasons, these differences may to some extent explain why length of hospital stay is longer in city hospitals than in rural ones.