18 qyGVyi3
Jurkat cells cultured in calcium-free RPMI-1640 medium (Gibco BRL; number {^>
m3
22300-107) containing calcium-free 10% FBS were triggered by anti-Fas IgM. The .]0B=w* Z
treated cells were harvested at the indicated time points and incubated with Fluo-3-AM at Q7UF
F
a final concentration of 1 micromolar for 30 min at 37°C (Scoltock et al., 2000). The &N GYV
labeled cells (50,000 cells per treatment) were then analyzed by exciting the cells at 488 z"bgtlfb8
nm and examining the fluorescence emission of Fluo 3 at 530 nm with a FACS Scan, n?_!gqK
(Becton Dickinson). A one micromolar concentration of LPA was used as a positive nzE4P3 C+
control for Ca2+ induction. The data thus obtained was analyzed with the software Win x+l.04a@
MDI 2.8 and represented as contour plots.The effect of chelating intracellular calcium on ]S<eO
6z
translocation of annexin I was studied by culturing Jurkat cells in the presence of 10 BRYhL|d~.
micromolar BAPTA-AM, with or without the addition of anti-Fas IgM. Cells were g*F?
harvested and fractionated as detailed above, and the S-100 fractions were assessed by +'Pl?QyH
immunoblotting for presence or absence of annexin I. o$oW-U
Mouse bone marrow transduction and transplantation. N[d*_KN.!
Retrovirus-mediated transduction of mouse bone marrow cells was done by published f;`pj`-k%
methods49. Prestimulated low-density bone marrow cells were infected with high-titer ]?S\So+
retrovirus supernatant on fibronectin-coated plates. Retrovirus supernatant was generated a_4Ny
in the phoenix-gp cells with a mouse stem cell virus–based retroviral vector coexpressing _Sy-&}c+
+
EGFP and HA–RhoH as described50. EGFP+ sorted cells were transplanted by `
4__X;
intravenous injection into the sublethally irradiated (300 rads with a 137Cs irradiator) 5p}Y6Lc\j
Rag2-/- recipient mice. At 9 weeks after transplantation, thymus, peripheral blood, bone *Uie{^p?
marrow, spleen and lymph nodes from each recipient mouse were collected for analysis uFOxb}a9v
of EGFP+ chimerism and hematopoietic lineage by flow cytometry. Expression of I-oI,c%+
HA–RhoH and HA–RhoHF73F83 in EGFP+ sorted thymocytes of recipient mice was 8o#*0d|
confirmed by immunoblot analysis. T`7;Rl'Q
Determination of renal morphology $D
+6=m[
Kidney slices were postfixed in buffered 2% OsO4, dehydrated, and embedded in an 0b+OB pqN
Araldite-EM bed 812 mixture. Large sections were cut perpendicular to the renal capsule, *gfx'$
containing cortex, and medulla. Thin (1 m) sections were analyzed in a blinded manner EGL1[7It`
for morphologic alterations, as previously detailed zOD5a=[1
Patient population Vd~{SS2>
Patients included in the study met the following criteria: (1) biopsy-proven IMN; (2) <h:> :%# k
creatinine clearance 30 ml per min per 1.73 m2; and (3) persistent proteinuria >5 g per -jnx0{/
24 h despite treatment with an HMG-CoA (3-hydroxy-3-methylglutaryl-coenzyme A) RX8$&z
reductase inhibitor, an ACEi, and/or ARB at maximal tolerated dose for at least 4 months. WL$Ee=
The Mayo Institutional Review Board and the Research Ethical Board, University Health 3XdN\xc
Network, University of Toronto approved the study protocol. All patients gave written 7;RhA5M
informed consent. Patients who had been on treatment with prednisone, cyclosporine, or N|K,{
p^li
19 ^IGutZov
mycophenolic mofetil within the last 4 months or alkylating agents within the last 6 A]"$O&l
months were not included in the study. Patients with active infection, diabetes, or a rq?:I:0
secondary cause of MN (for example, hepatitis B, systemic lupus erythematosus (SLE), BTjfzfO"
medications, malignancies) were also excluded. 0&T0Ls#
4
Treatment bg;NBoZd
At enrollment, a low-sodium (<4 g day-1) and low-protein (0.8 g per kg per day of x4|>HY<p?
high-quality protein) diet was recommended and patients were encouraged to maintain 7_C;-
the same diet throughout the duration of the study. All patients received a similar sKYb&2wJ
conservative treatment regimen that included loop diuretics to control edema, an 7sKN`
HMG-CoA reductase inhibitor, and an ACEi combined with an ARB if tolerated. wEL$QOu$
-Blockers and non-dihydropyridine calcium channel blockers, in that order, were added `l,=iy$
when required to control systolic blood pressures to <135 mm Hg in >75% of the
k_%2Ok
readings. Patients who after a minimum of 4 months of conservative therapy and {Ts@#V=:
maximized Ang II blockade had proteinuria >5 g per 24 h received two i.v. infusions of ~l"70\&
rituximab at a dose of 1000 mg on days 1 and 15. To minimize infusion reactions, vRa|lGeW
patients were premedicated with acetaminophen (1000 mg) and diphenhydramine 2`>/y
hydrochloride (50 mg) orally. In addition, methylprednisolone (100 mg, i.v.) was given 4GWt.+{J$
prior to the first rituximab infusion. B-cell depletion was defined as CD19+ count @s!9 T
<5 cells per l at any time and B-cell recovery was defined as CD19+ cell count ]:`q/iS&
>15 cells per l. Patients treated with rituximab, who at month 6 had proteinuria >3 g per e^kccz2f
24 h and in whom CD19+ B-cell counts had increased to >15 cells per l, received a q !\Ht2$b
second course of rituximab treatment following the same protocol described above. N),bhYS]
Follow-up }Kc[pp|9<
In all patients, clinical and laboratory parameters including complete blood counts, ^2E\{$J
electrolytes, serum albumin, B-cell flow cytometry for CD19+ B cells, serum }[ LME Z
immunoglobulin (IgG, IgM, IgA) levels, and a lipid panel were evaluated at study entry *(6vO{
and at months 1, 3, 6, 9, and 12. Creatinine clearance and protein and creatinine excretion A6ipA/_
in the urine were assessed by performing two consecutive 24-h urine collections at each h(]aP<49L
time point. Data were considered accurate when urinary creatinine excretion was nEu,1
consistent with a complete 24 h collection. The mean of the two measurements was *0zdI<Oe
considered for the analysis. The presence of HACAs was evaluated at baseline and at $;NxO0$
months 3, 6, 9, and 12. B:S/
?v
Method / Approach / Study/ Technique kKr|PFz
A discussion is presented of a problem-solving system X`bN/sI
To improve the efficiency of the method, the following approach may be applied.
i zJa`K
In order to an investigation was made to find the causes of the MmH[ 7R
Although large collections of rules and equations have been complied, none are generally _mVq9nBEf
accepted P0O5CaR
20 `Wq4k>J}*
This approach will be explained and discussed thoroughly in the body of the report. 2u 8z>/G
This can be accomplished by
fp!Ba
This algorithm to compute the total cost can be described step by step as follows: j3{8]D
The above preliminary analysis has provided important information YEfa8'7R
Various methods have been proposed for selecting an optimum... ph\KTLU
These concepts have been applied to o,l 3j|1
On the basis of the concept mentioned above, *Y!'3|T
This can be achieved by N&g9z{m7
In addition, tissues were stained for infiltrating lymphocytes (CD20 and CD3), and the IG@.W sM_
amount of interstitial fibrosis was quantified by histomorphometry. Formalin-fixed, 1haH2F^q3
paraffin-embedded sections were cut onto coated glass slides. Following heat-induced gBM6{48GF
antigen retrieval, sections were incubated at 20°C overnight with either anti-CD20 L3[r7 b
primary antibody or anti-CD3 primary antibody, both at 1:1000 dilution (Dako, Canada A[20i
c
Inc, Mississauga, Ontario, Canada). After rinsing all sections, pretreatment with 3% ^8dd
hydrogen peroxide was performed to prevent endogenous peroxidase activation. Sections .)Xyzd
were incubated with a secondary rabbit anti-mouse antibody linked with avidin–biotin c%p7?3Ry
complex. Sections were counterstained with hematoxylin and examined by light 7_t\wmvYp
microscopy. bvdAOvxChW
The HACA assay is a proprietary bridging enzyme-linked immunosorbent assay AQX~do\A
performed at Genentech Inc. that measures the antibody response to rituximab in human v:otR%yt
serum samples. &k'J5YHm8H
In all patients, clinical and laboratory parameters including complete blood counts, ^Mw>'*5^
electrolytes, serum albumin, B-cell flow cytometry for CD19+ B cells, serum xWkCP2$?P
immunoglobulin (IgG, IgM, IgA) levels, and a lipid panel were evaluated at study entry yZ7aH|Q81B
and at months 1, 3, 6, 9, and 12. 2c?
-_OCy;
This fact suggests that a new concept ut26sg{s(
This was accomplished by taking ... sg@)IEg</v
The preparatory stage is very time consuming process. aD
yHIh8
Test are performed for validity, completeness, and compatibility 5I #L
|+
There is little hope of achieving successful ... r\6 "mU
There has been an increasing awareness of the potential of using most ..so far made have E,"?RbG
not taken this approach, with the exception of w74)kIi
Only a few studies can be found. a.P7O!2Lp
It is a very tedious process to go through )b<-=VR
It is only when .. has been completed that .. may be effected MUo}Qi0K
The entire interpretation process is conducted in one's head. e[txJ*SuO
These approaches are sometimes very tedious. S $wx>715
Several techniques can be used |e+8Xz1>
A polynomial parametric model can be written as [the following]/[follows]: Y\e,#y
A xx model is constructed/formulated using xx. 5nq0#0Oc
A xx model represents an xx by its xx. sW/^82(dM
A process decision model captures the logic essential to DP E NYr
From the equation above, xx is equal to the summation of xx times the ... 5|l&` fv`
21 #BA=?7
The validity of a xx model can be checked using Euler's formula. 1~!
4
Given a model, one can mathematically determine whether ... or ... *5( h,s3&
Equations for xx need to be derived and implemented in the system. lq2Ah=FuN
A number of heuristic rules have been developed for 25xpq^Zw
Optimum .. techniques can be made more reliable by ... so that i:ar{ q
An algorithm based on the characteristic ... is used to determine <~:Lp:6 J
Euler's formula states the following: /S29
\^
The completed model should agree with the formula. W n*>h'R
For manufacturing purposes, a detailed and precise model of the object is necessary IooAXwOF
Engineering design models are very well defined; therefore, j^DoILw
To keep the domain narrow enough to be implementable, yet wide enough to be useful. KPGo*mY
Point of View Ap}^6_YXd
from an implementation standpoint,
BR1oE3in
From the point of view of this application, 9b`J2_ ]k
From this point of view, Zadeh suggested an inference rule named xxx (CRI for short).
w
7Do#Cv
Information is the meaningful interpretation and correlation of some aggregation of data HX1RA5O
in order to allow one to make decisions. b&k !DeE
From a practical point of view, the computational aspects of an FLC require a x5q5<-#
simplification of the fuzzy control algorithm.
NYwE=b~I
The use of a hammer to insert screws, although partly effective, tends to distort, destroy, Xo[={2_
and generally defeat the purpose of using a screw [Kusiak AI Implications for CIM [a~|{~?8
p.129] WLr\ l29
Statistical analysis 3ZN
m ,{
Data were analyzed by one-way analysis of variance comparing the three conditions CBC0X}_`
(sham operation, ischemic AKI, and bilateral nephrectomy) at each time point. If HorFQ?8
significant F-statistic from analysis of variance existed, this test was followed by Dunnett JGH;&UYP
post hoc multiple comparison procedure with sham operation as the control group. For all b/N+X}VMN
other comparisons, Student's t-test was used. A P-value of <0.05 was considered as h
WvQh
statistically significant. sdq8wn
Values are expressed as means s.e.m. and significance was evaluated by Mann–Whitney 7TCY$RcF,I
U test using GraphPad Prism, version 4.0 software (GraphPad Software Inc., San Diego, _~f&wkc
CA, USA). hr9[$4'H
All values are expressed as means s.d. Statistical significance (defined as P<0.05) was mfHZGk[[
evaluated using analysis of variance and Bonferroni t-tests, and the two-tailed Pearson's
'UGkL;
test, where appropriate.
BD6oN
]
Data are expressed as mean s.d., median and interquartile range, or frequencies, as Ew>E]Ys
appropriate. Variables who deviated from the normal distribution (positively skewed) 4(5NHsvp
were log-transformed (log10) before the correlation study. 5VWyc9Q
Data are represented as means s.e.m. Student's t-test and multiple comparisons with t-test Dh=?Hzw
post hoc analysis of variance were used as indicated below, for the comparison of 4L(/Z}(
22 |l)z^V!
morphological, immunohistological and functional parameters. Statistical significance p8CDFLuV
was set at P<0.05. sem:"
The primary efficacy parameter was defined as change in urinary protein excretion from :q_(=EA
baseline (week 0) to 12 months after treatment. The 12-month changes were tested z;? 32K
against zero using the paired t-test. Secondary end points included 6-month changes in Ke0j8|
protein; the number with PR or CR at 6 or 12 months; and changes in glomerular 'CiV=&3/
filtration rate (GFR), serum albumin, and lipid profiles. Study sample size was based on 9os>k*
the desire to have 80% power to detect a drop in urinary protein of at least 2.0 g day-1. '1|FqQ\.
Assuming a two-sided hypothesis test performed at a significance level of 0.05 and an s.d. rr/0pa$
of urinary protein change of 2.5 g, it was determined that 15 patients were required.2 I"TFj$Pg
Definition of remission status is according to the criteria established by Cattran et al.30 b 4o`eR
CR was defined as proteinuria <0.3 g per 24 h, PR as proteinuria 3 g per 24 h, and a nksx|i l
>50% reduction in peak proteinuria and non-response as <50% reduction in peak 9F[k;U
w
proteinuria. Any patient reaching a CR or PR was considered a treatment success bb@@QzR
The statistical significance of differences for the mean values of cytokine concentration
A.0eeX{
and T cell proliferation was determined with Student's t-test. Differences with a P value UVND1XV^f
of less than 0.05 were considered significant. x+niY;Z E
Statistical significance was determined by Student's t-test. Data with a P value of less [wHGt?R
than 0.05 were considered significant.
|Nx7jGd:i
In vitro and in vivo experiments were analyzed by the two-tailed Student's t-test, with P #^|"dIZ_M
values less than 0.05 considered statistically significant. NGsG4y^g?z
The significance of differences among groups was determined by Student's t-test and BhJag L ^o
one-way analysis of variance (SigmaStat software; Jandel). 6x?3%0Km
Comparisons tWD5Yh>.?$
As well, the pros and cons of these representations from a process planning point of view 2`2S94'
will be discussed.
mU
d['Z
The method of using xx to implement xx described by Zadeh (1973) appeared more IdP"]Sv{<
suitable Pra,r9h,
As discussed [in the previous section]/[preciously],
nQy %av$
Relation d~ m,hCTe
We can not invert F' directly because it defines a many-to-one mapping. NT3Ti
?J,
The relationships appear very complicate E@uxEF
Lifting tasks involve complex and imprecise relationship between the task variables and ~57.0?IK
the human operator's characteristics. S.^/Cl;aj
These methods are based on the relationship between ... and ... m]XG7:}V0
The fundamental concept of a fuzzy rating language is that we can establish a relationship ,\2w+L5TD
among terms such as high, medium, and low, and then modify these relationships. )I/K-zj
This article will thus mention the latter as well as the former. 2m! T.$
The former two bear a close relation to a fuzzy Cartesian product. 1/hk3m(C
Importance vh29mzum
) oQsls9t
23 c
N]e{|
The emphasis is on an implementation of a general approach to rule based decision Imv]V6"D=
making. Qj:`[#3?2
Consideration / Attention LBCat=d<
Careful evaluation is necessary to ensure T/9`VB%N
Such a formulation does not change further considerations. u)NmjW
Considerable attention has been paid to pb`!_GmB
Attention should be paid to an important finding of this investigation. W.|r=
Caution should be exercised in this process to avoid ... z8n=\xL
Primary consideration is given to ... components, though others can be accommodated IRq@~vdt)
After ... has been defined by ..., a carefully analysis is carried out/performed to determine i,3[0*ge
A number of factors such as ...need to be taken into consideration before making the 8M5a&