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= e;code = 101 char = r;code = 114 char = a;code = 97 char = t;code = 116 char = i;code = 105 char = v;code = 118 char = e;code = 101 char = ;code = 32 char = f;code = 102 char = a;code = 97 char = c;code = 99 char = t;code = 116 char = o;code = 111 char = r;code = 114 char = ;code = 32 char = (;code = 40 char = A;code = 65 char = P;code = 80 char = F;code = 70 char = );code = 41 char = ;code = 32 char = i;code = 105 char = s;code = 115 char = ;code = 32 char = a;code = 97 char = ;code = 32 char = s;code = 115 char = m;code = 109 char = a;code = 97 char = l;code = 108 char = l;code = 108 char = ;code = 32 char = g;code = 103 char = l;code = 108 char = y;code = 121 char = c;code = 99 char = o;code = 111 char = p;code = 112 char = r;code = 114 char = o;code = 111 char = t;code = 116 char = e;code = 101 char = i;code = 105 char = n;code = 110 char = ;code = 32 char = m;code = 109 char = a;code = 97 char = d;code = 100 char = e;code = 101 char = ;code = 32 char = s;code = 115 char = p;code = 112 char = e;code = 101 char = c;code = 99 char = i;code = 105 char = f;code = 102 char = i;code = 105 char = c;code = 99 char = a;code = 97 char = l;code = 108 char = l;code = 108 char = y;code = 121 char = ;code = 32 char = b;code = 98 char = y;code = 121 char = ;code = 32 char = b;code = 98 char = l;code = 108 char = a;code = 97 char = d;code = 100 char = d;code = 100 char = e;code = 101 char = r;code = 114 char = ;code = 32 char = e;code = 101 char = p;code = 112 char = i;code = 105 char = t;code = 116 char = h;code = 104 char = e;code = 101 char = l;code = 108 char = i;code = 105 char = a;code = 97 char = l;code = 108 char = ;code = 32 char = c;code = 99 char = e;code = 101 char = l;code = 108 char = l;code = 108 char = s;code = 115 char = ;code = 32 char = i;code = 105 char = n;code = 110 char = ;code = 32 char = p;code = 112 char = a;code = 97 char = t;code = 116 char = i;code = 105 char = e;code = 101 char = n;code = 110 char = t;code = 116 char = s;code = 115 char = ;code = 32 char = w;code = 119 char = i;code = 105 char = t;code = 116 char = h;code = 104 char = ;code = 32 char = I;code = 73 char = C;code = 67 char = ;code = 32 char = t;code = 116 char = h;code = 104 char = a;code = 97 char = t;code = 116 char = ;code = 32 char = i;code = 105 char = n;code = 110 char = d;code = 100 char = u;code = 117 char = c;code = 99 char = e;code = 101 char = s;code = 115 char = ;code = 32 char = c;code = 99 char = h;code = 104 char = a;code = 97 char = n;code = 110 char = g;code = 103 char = e;code = 101 char = s;code = 115 char = ;code = 32 char = i;code = 105 char = n;code = 110 char = ;code = 32 char = e;code = 101 char = x;code = 120 char = p;code = 112 char = r;code = 114 char = e;code = 101 char = s;code = 115 char = s;code = 115 char = i;code = 105 char = o;code = 111 char = n;code = 110 char = ;code = 32 char = o;code = 111 char = f;code = 102 char = ;code = 32 char = c;code = 99 char = e;code = 101 char = r;code = 114 char = t;code = 116 char = a;code = 97 char = i;code = 105 char = n;code = 110 char = ;code = 32 char = e;code = 101 char = p;code = 112 char = i;code = 105 char = t;code = 116 char = h;code = 104 char = e;code = 101 char = l;code = 108 char = i;code = 105 char = a;code = 97 char = l;code = 108 char = ;code = 32 char = c;code = 99 char = e;code = 101 char = l;code = 108 char = l;code = 108 char = ;code = 32 char = p;code = 112 char = r;code = 114 char = o;code = 111 char = t;code = 116 char = e;code = 101 char = i;code = 105 char = n;code = 110 char = s;code = 115 char = ;code = 32 char = a;code = 97 char = n;code = 110 char = d;code = 100 char = ;code = 32 char = p;code = 112 char = r;code = 114 char = o;code = 111 char = f;code = 102 char = o;code = 111 char = u;code = 117 char = n;code = 110 char = d;code = 100 char = l;code = 108 char = y;code = 121 char = ;code = 32 char = i;code = 105 char = n;code = 110 char = h;code = 104 char = i;code = 105 char = b;code = 98 char = i;code = 105 char = t;code = 116 char = s;code = 115 char = ;code = 32 char = c;code = 99 char = e;code = 101 char = l;code = 108 char = l;code = 108 char = ;code = 32 char = g;code = 103 char = r;code = 114 char = o;code = 111 char = w;code = 119 char = t;code = 116 char = h;code = 104 char = .;code = 46 char = ;code = 32 char = T;code = 84 char = h;code = 104 char = e;code = 101 char = r;code = 114 char = e;code = 101 char = f;code = 102 char = o;code = 111 char = r;code = 114 char = e;code = 101 char = ,;code = 44 char = ;code = 32 char = w;code = 119 char = e;code = 101 char = ;code = 32 char = c;code = 99 char = o;code = 111 char = n;code = 110 char = f;code = 102 char = i;code = 105 char = r;code = 114 char = m;code = 109 char = e;code = 101 char = d;code = 100 char = ;code = 32 char = t;code = 116 char = h;code = 104 char = e;code = 101 char = ;code = 32 char = i;code = 105 char = n;code = 110 char = c;code = 99 char = r;code = 114 char = e;code = 101 char = a;code = 97 char = s;code = 115 char = e;code = 101 char = d;code = 100 char = ;code = 32 char = p;code = 112 char = e;code = 101 char = r;code = 114 char = m;code = 109 char = e;code = 101 char = a;code = 97 char = b;code = 98 char = i;code = 105 char = l;code = 108 char = i;code = 105 char = t;code = 116 char = y;code = 121 char = ;code = 32 char = a;code = 97 char = n;code = 110 char = d;code = 100 char = ;code = 32 char = d;code = 100 char = e;code = 101 char = c;code = 99 char = r;code = 114 char = e;code = 101 char = a;code = 97 char = s;code = 115 char = e;code = 101 char = d;code = 100 char = ;code = 32 char = t;code = 116 char = i;code = 105 char = g;code = 103 char = h;code = 104 char = t;code = 116 char = ;code = 32 char = j;code = 106 char = u;code = 117 char = n;code = 110 char = c;code = 99 char = t;code = 116 char = i;code = 105 char = o;code = 111 char = n;code = 110 char = ;code = 32 char = f;code = 102 char = o;code = 111 char = r;code = 114 char = m;code = 109 char = a;code = 97 char = t;code = 116 char = i;code = 105 char = o;code = 111 char = n;code = 110 char = ;code = 32 char = o;code = 111 char = f;code = 102 char = ;code = 32 char = b;code = 98 char = l;code = 108 char = a;code = 97 char = d;code = 100 char = d;code = 100 char = e;code = 101 char = r;code = 114 char = ;code = 32 char = e;code = 101 char = p;code = 112 char = i;code = 105 char = t;code = 116 char = h;code = 104 char = e;code = 101 char = l;code = 108 char = i;code = 105 char = a;code = 97 char = l;code = 108 char = ;code = 32 char = c;code = 99 char = e;code = 101 char = l;code = 108 char = l;code = 108 char = ;code = 32 char = m;code = 109 char = o;code = 111 char = n;code = 110 char = o;code = 111 char = l;code = 108 char = a;code = 97 char = y;code = 121 char = e;code = 101 char = r;code = 114 char = s;code = 115 char = ;code = 32 char = g;code = 103 char = r;code = 114 char = o;code = 111 char = w;code = 119 char = n;code = 110 char = ;code = 32 char = f;code = 102 char = r;code = 114 char = o;code = 111 char = m;code = 109 char = ;code = 32 char = b;code = 98 char = i;code = 105 char = o;code = 111 char = p;code = 112 char = s;code = 115 char = i;code = 105 char = e;code = 101 char = s;code = 115 char = ;code = 32 char = i;code = 105 char = n;code = 110 char = ;code = 32 char = p;code = 112 char = a;code = 97 char = t;code = 116 char = i;code = 105 char = e;code = 101 char = n;code = 110 char = t;code = 116 char = s;code = 115 char = ;code = 32 char = w;code = 119 char = i;code = 105 char = t;code = 116 char = h;code = 104 char = ;code = 32 char = I;code = 73 char = C;code = 67 char = ;code = 32 char = c;code = 99 char = o;code = 111 char = m;code = 109 char = p;code = 112 char = a;code = 97 char = r;code = 114 char = e;code = 101 char = d;code = 100 char = ;code = 32 char = t;code = 116 char = o;code = 111 char = ;code = 32 char = c;code = 99 char = e;code = 101 char = l;code = 108 char = l;code = 108 char = s;code = 115 char = ;code = 32 char = f;code = 102 char = r;code = 114 char = o;code = 111 char = m;code = 109 char = ;code = 32 char = n;code = 110 char = o;code = 111 char = r;code = 114 char = m;code = 109 char = a;code = 97 char = l;code = 108 char = ;code = 32 char = c;code = 99 char = o;code = 111 char = n;code = 110 char = t;code = 116 char = r;code = 114 char = o;code = 111 char = l;code = 108 char = s;code = 115 char = .;code = 46 char = ;code = 32 char = W;code = 87 char = e;code = 101 char = ;code = 32 char = t;code = 116 char = h;code = 104 char = e;code = 101 char = n;code = 110 char = ;code = 32 char = d;code = 100 char = e;code = 101 char = t;code = 116 char = e;code = 101 char = r;code = 114 char = m;code = 109 char = i;code = 105 char = n;code = 110 char = e;code = 101 char = d;code = 100 char = ;code = 32 char = t;code = 116 char = h;code = 104 char = e;code = 101 char = ;code = 32 char = e;code = 101 char = f;code = 102 char = f;code = 102 char = e;code = 101 char = c;code = 99 char = t;code = 116 char = ;code = 32 char = o;code = 111 char = f;code = 102 char = ;code = 32 char = A;code = 65 char = P;code = 80 char = F;code = 70 char = ;code = 32 char = o;code = 111 char = n;code = 110 char = ;code = 32 char = t;code = 116 char = h;code = 104 char = e;code = 101 char = ;code = 32 char = p;code = 112 char = e;code = 101 char = r;code = 114 char = m;code = 109 char = e;code = 101 char = a;code = 97 char = b;code = 98 char = i;code = 105 char = l;code = 108 char = i;code = 105 char = t;code = 116 char = y;code = 121 char = ;code = 32 char = o;code = 111 char = f;code = 102 char = ;code = 32 char = n;code = 110 char = o;code = 111 char = r;code = 114 char = m;code = 109 char = a;code = 97 char = l;code = 108 char = ;code = 32 char = b;code = 98 char = l;code = 108 char = a;code = 97 char = d;code = 100 char = d;code = 100 char = e;code = 101 char = r;code = 114 char = ;code = 32 char = e;code = 101 char = p;code = 112 char = i;code = 105 char = t;code = 116 char = h;code = 104 char = e;code = 101 char = l;code = 108 char = i;code = 105 char = a;code = 97 char = l;code = 108 char = ;code = 32 char = c;code = 99 char = e;code = 101 char = l;code = 108 char = l;code = 108 char = ;code = 32 char = m;code = 109 char = o;code = 111 char = n;code = 110 char = o;code = 111 char = l;code = 108 char = a;code = 97 char = y;code = 121 char = e;code = 101 char = r;code = 114 char = s;code = 115 char = ;code = 32 char = a;code = 97 char = n;code = 110 char = d;code = 100 char = ;code = 32 char = t;code = 116 char = h;code = 104 char = e;code = 101 char = ;code = 32 char = e;code = 101 char = x;code = 120 char = p;code = 112 char = r;code = 114 char = e;code = 101 char = s;code = 115 char = s;code = 115 char = i;code = 105 char = o;code = 111 char = n;code = 110 char = ;code = 32 char = o;code = 111 char = f;code = 102 char = ;code = 32 char = t;code = 116 char = i;code = 105 char = g;code = 103 char = h;code = 104 char = t;code = 116 char = ;code = 32 char = j;code = 106 char = u;code = 117 char = n;code = 110 char = c;code = 99 char = t;code = 116 char = i;code = 105 char = o;code = 111 char = n;code = 110 char = ;code = 32 char = p;code = 112 char = r;code = 114 char = o;code = 111 char = t;code = 116 char = e;code = 101 char = i;code = 105 char = n;code = 110 char = s;code = 115 char = .;code = 46 char = <;code = 60 char = b;code = 98 char = r;code = 114 char = /;code = 47 char = >;code = 62 char = <;code = 60 char = b;code = 98 char = r;code = 114 char = /;code = 47 char = >;code = 62 char = <;code = 60 char = b;code = 98 char = >;code = 62 char = M;code = 77 char = A;code = 65 char = T;code = 84 char = E;code = 69 char = R;code = 82 char = I;code = 73 char = A;code = 65 char = L;code = 76 char = S;code = 83 char = ;code = 32 char = A;code = 65 char = N;code = 78 char = D;code = 68 char = ;code = 32 char = M;code = 77 char = E;code = 69 char = T;code = 84 char = H;code = 72 char = O;code = 79 char = D;code = 68 char = S;code = 83 char = ;code = 32 char = :;code = 58 char = ;code = 32 char = <;code = 60 char = /;code = 47 char = b;code = 98 char = >;code = 62 char = P;code = 80 char = e;code = 101 char = r;code = 114 char = m;code = 109 char = e;code = 101 char = a;code = 97 char = b;code = 98 char = i;code = 105 char = l;code = 108 char = i;code = 105 char = t;code = 116 char = y;code = 121 char = ;code = 32 char = w;code = 119 char = a;code = 97 char = s;code = 115 char = ;code = 32 char = d;code = 100 char = e;code = 101 char = t;code = 116 char = e;code = 101 char = r;code = 114 char = m;code = 109 char = i;code = 105 char = n;code = 110 char = e;code = 101 char = d;code = 100 char = ;code = 32 char = b;code = 98 char = y;code = 121 char = ;code = 32 char = m;code = 109 char = e;code = 101 char = a;code = 97 char = s;code = 115 char = u;code = 117 char = r;code = 114 char = i;code = 105 char = n;code = 110 char = g;code = 103 char = ;code = 32 char = t;code = 116 char = h;code = 104 char = e;code = 101 char = ;code = 32 char = C;code = 67 char = -;code = 45 char = m;code = 109 char = a;code = 97 char = 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;code = 32 char = m;code = 109 char = o;code = 111 char = n;code = 110 char = o;code = 111 char = l;code = 108 char = a;code = 97 char = y;code = 121 char = e;code = 101 char = r;code = 114 char = s;code = 115 char = ;code = 32 char = o;code = 111 char = n;code = 110 char = ;code = 32 char = T;code = 84 char = r;code = 114 char = a;code = 97 char = n;code = 110 char = s;code = 115 char = w;code = 119 char = e;code = 101 char = l;code = 108 char = l;code = 108 char = ;code = 32 char = c;code = 99 char = u;code = 117 char = l;code = 108 char = t;code = 116 char = u;code = 117 char = r;code = 114 char = e;code = 101 char = ;code = 32 char = p;code = 112 char = l;code = 108 char = a;code = 97 char = t;code = 116 char = e;code = 101 char = s;code = 115 char = ;code = 32 char = (;code = 40 char = C;code = 67 char = o;code = 111 char = r;code = 114 char = n;code = 110 char = i;code = 105 char = n;code = 110 char = g;code = 103 char = ,;code = 44 char = ;code = 32 char = C;code = 67 char = o;code = 111 char = r;code = 114 char = n;code = 110 char = i;code = 105 char = n;code = 110 char = g;code = 103 char = ,;code = 44 char = ;code = 32 char = N;code = 78 char = e;code = 101 char = w;code = 119 char = ;code = 32 char = Y;code = 89 char = o;code = 111 char = r;code = 114 char = k;code = 107 char = );code = 41 char = .;code = 46 char = ;code = 32 char = T;code = 84 char = i;code = 105 char = g;code = 103 char = h;code = 104 char = t;code = 116 char = ;code = 32 char = j;code = 106 char = u;code = 117 char = n;code = 110 char = c;code = 99 char = t;code = 116 char = i;code = 105 char = o;code = 111 char = n;code = 110 char = ;code = 32 char = f;code = 102 char = o;code = 111 char = r;code = 114 char = m;code = 109 char = a;code = 97 char = t;code = 116 char = i;code = 105 char = o;code = 111 char = n;code = 110 char = ;code = 32 char = w;code = 119 char = a;code = 97 char = s;code = 115 char = ;code = 32 char = a;code = 97 char = s;code = 115 char = s;code = 115 char = e;code = 101 char = s;code = 115 char = s;code = 115 char = e;code = 101 char = d;code = 100 char = ;code = 32 char = b;code = 98 char = y;code = 121 char = ;code = 32 char = i;code = 105 char = m;code = 109 char = m;code = 109 char = u;code = 117 char = n;code = 110 char = o;code = 111 char = f;code = 102 char = l;code = 108 char = u;code = 117 char = o;code = 111 char = r;code = 114 char = e;code = 101 char = s;code = 115 char = c;code = 99 char = e;code = 101 char = n;code = 110 char = c;code = 99 char = e;code = 101 char = ;code = 32 char = m;code = 109 char = i;code = 105 char = c;code = 99 char = r;code = 114 char = o;code = 111 char = s;code = 115 char = c;code = 99 char = o;code = 111 char = p;code = 112 char = y;code = 121 char = ;code = 32 char = a;code = 97 char = n;code = 110 char = d;code = 100 char = ;code = 32 char = t;code = 116 char = h;code = 104 char = e;code = 101 char = ;code = 32 char = e;code = 101 char = x;code = 120 char = p;code = 112 char = r;code = 114 char = e;code = 101 char = s;code = 115 char = s;code = 115 char = i;code = 105 char = o;code = 111 char = n;code = 110 char = ;code = 32 char = o;code = 111 char = f;code = 102 char = ;code = 32 char = s;code = 115 char = p;code = 112 char = e;code = 101 char = c;code = 99 char = i;code = 105 char = f;code = 102 char = i;code = 105 char = c;code = 99 char = ;code = 32 char = p;code = 112 char = r;code = 114 char = o;code = 111 char = t;code = 116 char = e;code = 101 char = i;code = 105 char = n;code = 110 char = s;code = 115 char = ;code = 32 char = w;code = 119 char = a;code = 97 char = s;code = 115 char = ;code = 32 char = d;code = 100 char = e;code = 101 char = t;code = 116 char = e;code = 101 char = r;code = 114 char = m;code = 109 char = i;code = 105 char = n;code = 110 char = e;code = 101 char = d;code = 100 char = ;code = 32 char = b;code = 98 char = y;code = 121 char = ;code = 32 char = W;code = 87 char = e;code = 101 char = s;code = 115 char = t;code = 116 char = e;code = 101 char = r;code = 114 char = n;code = 110 char = ;code = 32 char = b;code = 98 char = l;code = 108 char = o;code = 111 char = t;code = 116 char = .;code = 46 char = <;code = 60 char = b;code = 98 char = r;code = 114 char = /;code = 47 char = >;code = 62 char = <;code = 60 char = b;code = 98 char = r;code = 114 char = /;code = 47 char = >;code = 62 char = <;code = 60 char = b;code = 98 char = >;code = 62 char = R;code = 82 char = E;code = 69 char = S;code = 83 char = U;code = 85 char = L;code = 76 char = T;code = 84 char = S;code = 83 char = ;code = 32 char = :;code = 58 char = ;code = 32 char = <;code = 60 char = /;code = 47 char = b;code = 98 char = >;code = 62 char = A;code = 65 char = P;code = 80 char = F;code = 70 char = ;code = 32 char = t;code = 116 char = r;code = 114 char = e;code = 101 char = a;code = 97 char = t;code = 116 char = m;code = 109 char = e;code = 101 char = n;code = 110 char = t;code = 116 char = ;code = 32 char = c;code = 99 char = a;code = 97 char = u;code = 117 char = s;code = 115 char = e;code = 101 char = d;code = 100 char = ;code = 32 char = s;code = 115 char = i;code = 105 char = g;code = 103 char = n;code = 110 char = i;code = 105 char = f;code = 102 char = i;code = 105 char = c;code = 99 char = a;code = 97 char = n;code = 110 char = t;code = 116 char = ;code = 32 char = i;code = 105 char = n;code = 110 char = c;code = 99 char = r;code = 114 char = e;code = 101 char = a;code = 97 char = s;code = 115 char = e;code = 101 char = s;code = 115 char = ;code = 32 char = i;code = 105 char = n;code = 110 char = ;code = 32 char = t;code = 116 char = h;code = 104 char = e;code = 101 char = ;code = 32 char = p;code = 112 char = a;code = 97 char = r;code = 114 char = a;code = 97 char = c;code = 99 char = e;code = 101 char = l;code = 108 char = l;code = 108 char = u;code = 117 char = l;code = 108 char = a;code = 97 char = r;code = 114 char = ;code = 32 char = p;code = 112 char = e;code = 101 char = r;code = 114 char = m;code = 109 char = e;code = 101 char = a;code = 97 char = b;code = 98 char = i;code = 105 char = l;code = 108 char = i;code = 105 char = t;code = 116 char = y;code = 121 char = ;code = 32 char = o;code = 111 char = f;code = 102 char = ;code = 32 char = n;code = 110 char = o;code = 111 char = r;code = 114 char = m;code = 109 char = a;code = 97 char = l;code = 108 char = ;code = 32 char = b;code = 98 char = l;code = 108 char = a;code = 97 char = d;code = 100 char = d;code = 100 char = e;code = 101 char = r;code = 114 char = ;code = 32 char = e;code = 101 char = p;code = 112 char = i;code = 105 char = t;code = 116 char = h;code = 104 char = e;code = 101 char = l;code = 108 char = i;code = 105 char = a;code = 97 char = l;code = 108 char = ;code = 32 char = c;code = 99 char = e;code = 101 char = l;code = 108 char = l;code = 108 char = ;code = 32 char = m;code = 109 char = o;code = 111 char = n;code = 110 char = o;code = 111 char = l;code = 108 char = a;code = 97 char = y;code = 121 char = e;code = 101 char = r;code = 114 char = s;code = 115 char = ;code = 32 char = a;code = 97 char = n;code = 110 char = d;code = 100 char = ;code = 32 char = t;code = 116 char = h;code = 104 char = e;code = 101 char = ;code = 32 char = a;code = 97 char = t;code = 116 char = t;code = 116 char = e;code = 101 char = n;code = 110 char = u;code = 117 char = a;code = 97 char = t;code = 116 char = i;code = 105 char = o;code = 111 char = n;code = 110 char = ;code = 32 char = o;code = 111 char = f;code = 102 char = ;code = 32 char = t;code = 116 char = i;code = 105 char = g;code = 103 char = h;code = 104 char = t;code = 116 char = ;code = 32 char = j;code = 106 char = u;code = 117 char = n;code = 110 char = c;code = 99 char = t;code = 116 char = i;code = 105 char = o;code = 111 char = n;code = 110 char = s;code = 115 char = ;code = 32 char = c;code = 99 char = o;code = 111 char = m;code = 109 char = p;code = 112 char = a;code = 97 char = r;code = 114 char = e;code = 101 char = d;code = 100 char = ;code = 32 char = t;code = 116 char = o;code = 111 char = ;code = 32 char = m;code = 109 char = o;code = 111 char = c;code = 99 char = k;code = 107 char = ;code = 32 char = A;code = 65 char = P;code = 80 char = F;code = 70 char = ,;code = 44 char = ;code = 32 char = s;code = 115 char = i;code = 105 char = m;code = 109 char = i;code = 105 char = l;code = 108 char = a;code = 97 char = r;code = 114 char = ;code = 32 char = t;code = 116 char = o;code = 111 char = ;code = 32 char = c;code = 99 char = h;code = 104 char = a;code = 97 char = n;code = 110 char = g;code = 103 char = e;code = 101 char = s;code = 115 char = ;code = 32 char = s;code = 115 char = e;code = 101 char = e;code = 101 char = n;code = 110 char = ;code = 32 char = i;code = 105 char = n;code = 110 char = ;code = 32 char = I;code = 73 char = C;code = 67 char = ;code = 32 char = c;code = 99 char = e;code = 101 char = l;code = 108 char = l;code = 108 char = s;code = 115 char = .;code = 46 char = ;code = 32 char = A;code = 65 char = P;code = 80 char = F;code = 70 char = ;code = 32 char = t;code = 116 char = r;code = 114 char = e;code = 101 char = a;code = 97 char = t;code = 116 char = m;code = 109 char = e;code = 101 char = n;code = 110 char = t;code = 116 char = ;code = 32 char = a;code = 97 char = l;code = 108 char = s;code = 115 char = o;code = 111 char = ;code = 32 char = d;code = 100 char = e;code = 101 char = c;code = 99 char = r;code = 114 char = e;code = 101 char = a;code = 97 char = s;code = 115 char = e;code = 101 char = d;code = 100 char = ;code = 32 char = e;code = 101 char = x;code = 120 char = p;code = 112 char = r;code = 114 char = e;code = 101 char = s;code = 115 char = s;code = 115 char = i;code = 105 char = o;code = 111 char = n;code = 110 char = ;code = 32 char = o;code = 111 char = f;code = 102 char = ;code = 32 char = t;code = 116 char = h;code = 104 char = e;code = 101 char = ;code = 32 char = t;code = 116 char = i;code = 105 char = g;code = 103 char = h;code = 104 char = t;code = 116 char = ;code = 32 char = j;code = 106 char = u;code = 117 char = n;code = 110 char = c;code = 99 char = t;code = 116 char = i;code = 105 char = o;code = 111 char = n;code = 110 char = ;code = 32 char = p;code = 112 char = r;code = 114 char = o;code = 111 char = t;code = 116 char = e;code = 101 char = i;code = 105 char = n;code = 110 char = s;code = 115 char = ;code = 32 char = z;code = 122 char = o;code = 111 char = n;code = 110 char = u;code = 117 char = l;code = 108 char = a;code = 97 char = ;code = 32 char = o;code = 111 char = c;code = 99 char = c;code = 99 char = l;code = 108 char = u;code = 117 char = d;code = 100 char = e;code = 101 char = n;code = 110 char = s;code = 115 char = -;code = 45 char = 1;code = 49 char = ;code = 32 char = a;code = 97 char = n;code = 110 char = d;code = 100 char = ;code = 32 char = o;code = 111 char = c;code = 99 char = c;code = 99 char = l;code = 108 char = u;code = 117 char = d;code = 100 char = i;code = 105 char = n;code = 110 char = .;code = 46 char = <;code = 60 char = b;code = 98 char = r;code = 114 char = /;code = 47 char = >;code = 62 char = <;code = 60 char = b;code = 98 char = r;code = 114 char = /;code = 47 char = >;code = 62 char = <;code = 60 char = b;code = 98 char = >;code = 62 char = C;code = 67 char = O;code = 79 char = N;code = 78 char = C;code = 67 char = L;code = 76 char = U;code = 85 char = S;code = 83 char = I;code = 73 char = O;code = 79 char = N;code = 78 char = S;code = 83 char = ;code = 32 char = :;code = 58 char = ;code = 32 char = <;code = 60 char = /;code = 47 char = b;code = 98 char = >;code = 62 char = B;code = 66 char = e;code = 101 char = c;code = 99 char = a;code = 97 char = u;code = 117 char = s;code = 115 char = e;code = 101 char = ;code = 32 char = o;code = 111 char = f;code = 102 char = ;code = 32 char = i;code = 105 char = t;code = 116 char = s;code = 115 char = ;code = 32 char = a;code = 97 char = p;code = 112 char = p;code = 112 char = a;code = 97 char = r;code = 114 char = e;code = 101 char = n;code = 110 char = t;code = 116 char = ;code = 32 char = e;code = 101 char = f;code = 102 char = f;code = 102 char = e;code = 101 char = c;code = 99 char = t;code = 116 char = s;code = 115 char = ;code = 32 char = o;code = 111 char = n;code = 110 char = ;code = 32 char = b;code = 98 char = l;code = 108 char = a;code = 97 char = d;code = 100 char = d;code = 100 char = e;code = 101 char = r;code = 114 char = ;code = 32 char = e;code = 101 char = p;code = 112 char = i;code = 105 char = t;code = 116 char = h;code = 104 char = e;code = 101 char = l;code = 108 char = i;code = 105 char = a;code = 97 char = l;code = 108 char = ;code = 32 char = c;code = 99 char = e;code = 101 char = l;code = 108 char = l;code = 108 char = ;code = 32 char = t;code = 116 char = i;code = 105 char = g;code = 103 char = h;code = 104 char = t;code = 116 char = ;code = 32 char = j;code = 106 char = u;code = 117 char = n;code = 110 char = c;code = 99 char = t;code = 116 char = i;code = 105 char = o;code = 111 char = n;code = 110 char = s;code = 115 char = ;code = 32 char = a;code = 97 char = n;code = 110 char = d;code = 100 char = ;code = 32 char = p;code = 112 char = a;code = 97 char = r;code = 114 char = a;code = 97 char = c;code = 99 char = e;code = 101 char = l;code = 108 char = l;code = 108 char = u;code = 117 char = l;code = 108 char = a;code = 97 char = r;code = 114 char = ;code = 32 char = p;code = 112 char = e;code = 101 char = r;code = 114 char = m;code = 109 char = e;code = 101 char = a;code = 97 char = b;code = 98 char = i;code = 105 char = l;code = 108 char = i;code = 105 char = t;code = 116 char = y;code = 121 char = ;code = 32 char = i;code = 105 char = n;code = 110 char = ;code = 32 char = v;code = 118 char = i;code = 105 char = t;code = 116 char = r;code = 114 char = o;code = 111 char = ,;code = 44 char = ;code = 32 char = A;code = 65 char = P;code = 80 char = F;code = 70 char = ;code = 32 char = m;code = 109 char = a;code = 97 char = y;code = 121 char = ;code = 32 char = c;code = 99 char = o;code = 111 char = n;code = 110 char = t;code = 116 char = r;code = 114 char = i;code = 105 char = b;code = 98 char = u;code = 117 char = t;code = 116 char = e;code = 101 char = ;code = 32 char = t;code = 116 char = o;code = 111 char = ;code = 32 char = t;code = 116 char = h;code = 104 char = e;code = 101 char = ;code = 32 char = l;code = 108 char = e;code = 101 char = a;code = 97 char = k;code = 107 char = i;code = 105 char = n;code = 110 char = e;code = 101 char = s;code = 115 char = s;code = 115 char = ;code = 32 char = o;code = 111 char = f;code = 102 char = ;code = 32 char = t;code = 116 char = h;code = 104 char = e;code = 101 char = ;code = 32 char = b;code = 98 char = l;code = 108 char = a;code = 97 char = d;code = 100 char = d;code = 100 char = e;code = 101 char = r;code = 114 char = ;code = 32 char = e;code = 101 char = p;code = 112 char = i;code = 105 char = t;code = 116 char = h;code = 104 char = e;code = 101 char = l;code = 108 char = i;code = 105 char = a;code = 97 char = l;code = 108 char = ;code = 32 char = b;code = 98 char = a;code = 97 char = r;code = 114 char = r;code = 114 char = i;code = 105 char = e;code = 101 char = r;code = 114 char = ;code = 32 char = s;code = 115 char = e;code = 101 char = e;code = 101 char = n;code = 110 char = ;code = 32 char = i;code = 105 char = n;code = 110 char = ;code = 32 char = I;code = 73 char = C;code = 67 char = .;code = 46 result sql = PURPOSE : Previous reports have suggested that the bladder epithelial barrier may be compromised in interstitial cystitis (IC). Antiproliferative factor (APF) is a small glycoprotein made specifically by bladder epithelial cells in patients with IC that induces changes in expression of certain epithelial cell proteins and profoundly inhibits cell growth. Therefore, we confirmed the increased permeability and decreased tight junction formation of bladder epithelial cell monolayers grown from biopsies in patients with IC compared to cells from normal controls. We then determined the effect of APF on the permeability of normal bladder epithelial cell monolayers and the expression of tight junction proteins.MATERIALS AND METHODS : Permeability was determined by measuring the C-mannitol and H-inulin flux between cells in confluent monolayers on Transwell culture plates (Corning, Corning, New York). Tight junction formation was assessed by immunofluorescence microscopy and the expression of specific proteins was determined by Western blot.RESULTS : APF treatment caused significant increases in the paracellular permeability of normal bladder epithelial cell monolayers and the attenuation of tight junctions compared to mock APF, similar to changes seen in IC cells. APF treatment also decreased expression of the tight junction proteins zonula occludens-1 and occludin.CONCLUSIONS : Because of its apparent effects on bladder epithelial cell tight junctions and paracellular permeability in vitro, APF may contribute to the leakiness of the bladder epithelial barrier seen in IC.
Publication: J. Urol. 2005 Dec ;174(6)::2382-7.
Publication Date: 2005 Dec
Study Author(s): Zhang, Chen-Ou;Wang, Jian-Ying;Koch, Kristopher R;Keay, Susan;
Institution: Department of Pathology, University of Maryland School of Medicine, Baltimore, MD, USA.
Shortcut link to this study:
PURPOSE : Previous reports have suggested that the bladder epithelial barrier may be compromised in interstitial cystitis (IC). Antiproliferative factor (APF) is a small glycoprotein made specifically by bladder epithelial cells in patients with IC that induces changes in expression of certain epithelial cell proteins and profoundly inhibits cell growth. Therefore, we confirmed the increased permeability and decreased tight junction formation of bladder epithelial cell monolayers grown from biopsies in patients with IC compared to cells from normal controls. We then determined the effect of APF on the permeability of normal bladder epithelial cell monolayers and the expression of tight junction proteins.MATERIALS AND METHODS : Permeability was determined by measuring the C-mannitol and H-inulin flux between cells in confluent monolayers on Transwell culture plates (Corning, Corning, New York). Tight junction formation was assessed by immunofluorescence microscopy and the expression of specific proteins was determined by Western blot.RESULTS : APF treatment caused significant increases in the paracellular permeability of normal bladder epithelial cell monolayers and the attenuation of tight junctions compared to mock APF, similar to changes seen in IC cells. APF treatment also decreased expression of the tight junction proteins zonula occludens-1 and occludin.CONCLUSIONS : Because of its apparent effects on bladder epithelial cell tight junctions and paracellular permeability in vitro, APF may contribute to the leakiness of the bladder epithelial barrier seen in IC.
PMID: 16280852