12th Congress of the European Hematology ... - Haematologica

tinely by our stem cell lab prior to cell storage. Collection efficiency was
calculated as the ratio cells collected / cells processed. Effect of a variety
of parameters on collection efficiency was assessed, including: individual
Cobe Spectra machine used for collection; gender; length of procedure;
paediatric versus adult collection; average inlet flow speed (both
absolute and relative to total blood volume); allogeneic versus autologous
collection; and day of collection (for donors collected on several days running).
Results. There was a very strong linear correlation between cells
processed and cells collected (r=0.979), corresponding to a relatively constant
and predictable collection efficiency averaging out at 55%. Factors
appearing to influence collection efficiency positively included paediatric
procedures, and very slow absolute runspeeds (less than 25
mls/min), presumably in both cases due to longer PBSC dwell time in the
collection channel. Factors influencing collection efficiency negatively
included very short procedures (less than 150 minutes) and third or subsequent
day of collection (presumably due to collection on rapidly falling
peripheral CD34 counts). Factors which did not influence collection efficiency
significantly included: very long procedures (interestingly, there
was no evidence of stem cell depletion in procedures lasting 240 minutes
or more); gender; very fast runspeeds; runspeed relative to total blood
volume; and allogeneic versus autologous collection. Conclusions. Comparison
of average collection efficiencies between the five individual
Cobe Spectra machines was possible, yielding valuable information for
validation and process qualification of PBSC collection using the MNC
programme on these machines. Although there was some slight
machine-to-machine variability, this did not reach statistical significance.
The data were useful for two other specific quality purposes. Firstly, the
relative constancy of collection efficiency allows the predicted CD34 +
dose in the PBSC product to be calculated at the start of each collection
procedure, using the predicted end-run data provided by the machine
and the formula: Predicted dose = (5 x Peripheral CD34 + count/mL x
Final Inlet Flow in mls) / (Weight in kg x 10,000). (This formula includes
a correction for anticoagulant volume.) Secondly, the actual dose
achieved can be compared retrospectively against the predicted dose, and
(rare) collections where the actual dose achieved is less than half of the
predicted dose can then be investigated for quality purposes: in other
words, this provides a product specification for PBSC collections. Calculation
of predicted cell dose with a good degree of accuracy at the start of
a collection procedure is also useful to apheresis staff and to the patient
for planning purposes.
Figure 1. Correlation between Cells processed & Cells collected (all patients).
1043
EXPRESSION OF APOPTOSIS-REGULATING GENES BCL-2 AND BAX EVALUATED
BY QRT-PCR IN LYMPH NODES OF PATIENTS WITH HIGH GRADE NON-HODGKIN’S
LYMPHOMA
S. Woszczyk
Medical University of Silesia, KATOWICE, Poland
Background. Impairment of apoptosis is central to cancer development
(including lymphoma) and renders tumors refractory to cytotoxic therapy.
The balance between Bcl-2 and Bax is important for the induction
of programmed cell death; when Bcl-2 predominates, apoptosis is inhibited.
BCL-2 expression has been studied extensively in aggressive Non-
12 th Congress of the European Hematology Association
Hodgkin’s lymphomas, principally Diffuse Large B-cell Lymphoma
/DLBCL/, and has been correlated with worse prognosis. In contrast,
Bax expression and their clinical significance are not so well studied, the
results often being discordant. Moreover, most of the research conducted
so far has analyzed the activity of pro- and antiapoptotic genes in
peripheral blond mononuclears and little research has assessed this activity
in lymph nodes by means of mRNA analysis. Aims. The aim of study
was to evaluate the expression of apoptosis-regulating proteins Bcl-2
and Bax in the lymph nodes of 23 never-treated high grade Non-
Hodgkin’s lymphoma patients in different stages of the disease (15 of
these patients were diagnosed with DLBCL, 8 with the Mantle Cell Lymphoma-MCL;
the control group consisted of 7 patients with persistent
chronic lymphadenitis. Methods. The QRT-PCR method was employed
to assess the activity of Bcl-2 and Bax. Results. No difference in the expression
of Bcl-2 and Bax was found between MCL and the DLBCL groups
/p>0,05 U Mann-Whitney test/. The T-Student test, however, did show
a significantly higher expression level of Bcl-2 in the MCL group when
compared with the DLBCL group / p