scholarly journals Cortical cell populations from rabbit kidney isolated by free-flow electrophoresis: characterization by measurement of hormone-sensitive adenylate cyclase.

1982 ◽  
Vol 92 (2) ◽  
pp. 505-513 ◽  
Author(s):  
A Vandewalle ◽  
B Köpfer-Hobelsberger ◽  
H G Heidrich
Keyword(s):  
Alkaline Phosphatase ◽  
Parathyroid Hormone ◽  
Adenylate Cyclase ◽  
Fast Moving ◽  
Cortical Cell ◽  
Free Flow ◽  
Rabbit Kidney ◽  
Kidney Cortex ◽  
Cell Populations ◽  
Slow Moving

Free-flow electrophoresis allows the separation of different cell populations from a cell suspension isolated from rabbit kidney cortex after perfusion of the kidneys with a calcium-binder, followed by gentle mechanical treatment. After electrophoretic separation, analysis of the adenylate cyclase activities after stimulation by various hormones allows the precise determination of the origin of the cell populations with different electrophoretic mobilities. Adenylate cyclase from the slow-moving main cell population was only sensitive to parathyroid hormone. These cells had also high alkaline phosphatase content, further demonstrating their proximal origin. The various fast-moving cell populations had adenylate cyclase sensitive to isoproterenol and arginine vasopressin but were less sensitive to parathyroid hormone than the slow-moving cells. Their alkaline phosphatase content was also much lower. This indicates that these fast-moving cell populations originate from both the granulous segment of the distal tubule and from the collecting ducts. The adenylate cyclase activity and the cyclic AMP contents of isolated proximal cells maintained in culture medium were also investigated.

Separation of two bone cell populations from fetal rat calvaria and a study of their responses to parathyroid hormone and calcitonin

1983 ◽  
Vol 99 (3) ◽  
pp. 387-399 ◽  
Author(s):  
I. P. Braidman ◽  
D. C. Anderson ◽  
C. J. P. Jones ◽  
J. B. Weiss
Keyword(s):  
Alkaline Phosphatase ◽  
Parathyroid Hormone ◽  
Acid Phosphatase ◽  
B Cells ◽  
Adenylate Cyclase ◽  
Culture Medium ◽  
Cell Populations ◽  
C Cells ◽  
Type B ◽  
Type C

Bone cells released from perinatal rat calvaria by digestion with clostridial peptidase were separated into two distinct populations (designated types B and C) by equilibrium density centrifugation on a two-step gradient of Percoll. They were extensively characterized by light and electron microscopy and for behaviour in culture, acid and alkaline phosphatase activity, collagen synthesis, collagenase secretion and adenylate cyclase response to parathyroid hormone (PTH) and calcitonin. Type C cells were predominantly large with up to seven nuclei and an unusual cytoplasmic appearance in cytocentrifuge preparations. They did not proliferate in culture and we have established culture conditions which prevented their overgrowth by contaminating proliferative cells. In culture these cells had low alkaline and high acid phosphatase and high aryl sulphatase activity, and synthesized little collagen. In contrast type B cells were mostly smaller and many had irregular cytoplasmic projections. In culture they became polygonal in shape, proliferated rapidly, and reached confluence in 4–5 days. These were low in aryl sulphatase and acid phosphatase, high in alkaline phosphatase activity, and synthesized labelled collagen actively with [3H]proline and ascorbic acid included in the culture medium. The two cell populations were found to differ in culture in two important further respects. First, the type C cells showed an adenylate cyclase response to calcitonin but not to PTH, while the converse was true for type B cells; this was so over at least a 20-fold range of isobutylmethyl xanthine concentration. Secondly, type C cells in culture secreted an active collagenolytic enzyme. Type B cells secreted much lower levels of a predominantly latent collagenase which required activation by mersalyl. Co-culture of type C and type B cells led to a marked reduction in the content of active collagenase in the culture medium.

scholarly journals Regulation of adenylate cyclase activity by protein factors in rabbit kidney cortex

1979 ◽  
Vol 29 ◽  
pp. 87
Author(s):  
Tadaomi Takenawa ◽  
Katsutoshi Goto ◽  
Masaaki Kuroda ◽  
Tomoh Masaki
Keyword(s):  
Adenylate Cyclase ◽  
Cyclase Activity ◽  
Adenylate Cyclase Activity ◽  
Rabbit Kidney ◽  
Kidney Cortex

Indirect immunoselection of late distal cell populations from rabbit kidney cortex

1986 ◽  
Vol 250 (3) ◽  
pp. F386-F395 ◽  
Author(s):  
A. Vandewalle ◽  
M. Tauc ◽  
F. Cluzeaud ◽  
P. Ronco ◽  
F. Chatelet ◽  
...  
Keyword(s):  
Cell Suspension ◽  
Brush Border ◽  
Phosphoenolpyruvate Carboxykinase ◽  
High Yield ◽  
Rabbit Kidney ◽  
Kidney Cortex ◽  
Cell Populations ◽  
Cortical Cells ◽  
Rate Of Oxygen Consumption ◽  
Distal Cell

This study describes a method for the separation of distal cell populations based on the sequestration of proximal cells on immunoadsorbent columns (CNBr-activated Sepharose 6MB) bound with three brush-border monoclonal antibodies (S6-Mab). A high yield of isolated cell suspension from rabbit kidney cortex was prepared by mechanical dissociation after perfusion and incubation of the kidneys with 10(-3) M EDTA. The sequestration of the proximal cells was achieved in two sequential chromatographic steps. About 92% of the applied cells were first retained on an S6-Mab column after a 60-min stationary stage and the unbound cells were submitted by direct flow to a second S6-Mab column. In such conditions, 8 X 10(6) cells were recovered when starting with 331 X 10(6) cortical cells. The efficiency of the proximal cell depletion process was confirmed by an 80% decrease in brush-border enzymes, a very low phosphoenolpyruvate carboxykinase activity, and absence of cells bearing long microvilli, as ascertained by electron microscopy. This immunodepleted cell population presented the enzymatical characteristics of cells from the more distal segments. As compared with the initial cell suspension, these cells exhibited higher hexokinase (2.3 times), succinate dehydrogenase (1.5 times), and Na+-K+-ATPase (2.6 times) activities. In addition, adenylate cyclase activities remained sensitive to parathormone, arginine vasopressin, and isoproterenol. The functional capacity of these immunodepleted cells was assessed by an almost complete exclusion of eosin dye, a low Na+ and high K+ intracellular content, and a high respiratory rate of oxygen consumption. In conclusion, this immunoselective process makes it possible to obtain subpopulations of renal cortical cells possessing the main characteristics of the distal, connecting, and collecting cells for physiological and metabolic studies.

scholarly journals Binding of parathyroid hormone to bovine kidney-cortex plasma membranes

1973 ◽  
Vol 134 (4) ◽  
pp. 913-921 ◽  
Author(s):  
H. S. Sutcliffe ◽  
T. J. Martin ◽  
J. A. Eisman ◽  
R. Pilczyk
Keyword(s):  
Parathyroid Hormone ◽  
Adenylate Cyclase ◽  
Plasma Membranes ◽  
Specific Activity ◽  
Specific Binding ◽  
Rat Kidney ◽  
Kidney Cortex ◽  
Bovine Kidney ◽  
Hormone Sensitive ◽  
Chloramine T

1. Plasma membranes were purified from bovine kidney cortex, with a fourfold increase in specific activity of parathyroid hormone-sensitive adenylate cyclase over that in the crude homogenate. The membranes were characterized by enzyme studies. 2. Parathyroid hormone was labelled with 125I by an enzymic method and the labelled hormone shown to bind to the plasma membranes and to be specifically displaced by unlabelled hormone. Parathyroid hormone labelled by the chloramine-t procedure showed no specific binding. 75Se-labelled human parathyroid hormone, prepared in cell culture, also bound to the membranes. 3. Parathyroid hormone was shown to retain biological activity after iodination by the enzymic method, but no detectable activity remained after chloramine-t treatment. 4. High concentration of pig insulin inhibited binding of labelled parathyroid hormone to plasma membranes and partially inhibited the hormone-sensitive adenylate cyclase activity in a crude kidney-cortex preparation. 5. EDTA enhanced and Ca2+ inhibited binding of labelled parathyroid hormone to plasma membranes. 6. Whereas rat kidney homogenates were capable of degrading labelled parathyroid hormone to trichloroacetic acid-soluble fragments, neither crude homogenates nor purified membranes from bovine kidney showed this property. 7. Binding of parathyroid hormone is discussed in relation to metabolism and initial events in hormone action.

scholarly journals Parathyroid hormone acutely increases polyphosphoinositides of the rabbit kidney cortex by a cycloheximide-sensitive process.

1980 ◽  
Vol 65 (6) ◽  
pp. 1523-1526 ◽  
Author(s):  
R V Farese ◽  
P Bidot-López ◽  
A Sabir ◽  
J S Smith ◽  
B Schinbeckler ◽  
...  
Keyword(s):  
Parathyroid Hormone ◽  
Rabbit Kidney ◽  
Kidney Cortex
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