Significant pain relief with loading dose zoledronic acid in bone metastases is only seen in patients with elevated initial serum C telopeptide (CTX).

In symptomatic bone metastases with significant pain, refractory to standard analgesics and radiotherapy, loading dose zoledronic acid (ZA) represents a simple and nontoxic treatment to obtain significant pain relief in a very short time. Its analgesic effect is limited to patients with massive osteoclast activation with high initial serum C-telopeptide (CTX). The pain reduction is proportionally correlated with the reduction of CTX.

Osmotic effects of dilution on erythrocytes after freezing and thawing in glycerol-containing buffer.

Red blood cells frozen in 1.7 M and particularly in 2.2 M of glycerol retain a high degree of integrity upon thawing as long as the dilution procedure of the cryoprotectant is slow and preferentially compensated by the addition of sorbitol. As the nonpenetrating cryoprotectant sorbitol induces initial cell shrinkage, cell swelling upon dilution of the cryoprotectants may not lead to hemolysis. However, rapid dilution of glycerol even with buffer containing up to 0.50 M of sorbitol cannot be achieved without provoking considerable hemolysis. Due to the relative slow rate at which glycerol leaves the cells, membrane damage to the younger cell populations remains considerable and is even more pronounced in the older cell groups. The dramatic osmotic changes occurring during the dilution process lead to the formation of aberrant cell populations as demonstrated by the red cell size frequency distribution curves.

The prevention of erythrocyte swelling upon dilution after freezing and thawing.

Cellular swelling of erythrocytes exposed to Me2SO during freezing and thawing may lead to hemolysis upon dilution of the cryoprotectant with pure electrolyte buffer. Excessive cell swelling is effectively avoided by exposing the RBC to the nonpenetrating sorbitol after thawing and before dilution. Due to the initial reduction in volume by sorbitol, cell swelling upon dilution may not cause hemolysis particularly with concentrations of 0.05 to 0.15 M of sorbitol in the diluting electrolyte buffer. Membrane damage incurred during freezing and thawing is particularly pronounced with the older red cell population, while the younger population membrane integrity can be preserved to an optimal degree.

The effects of cryopreservation on membrane integrity, membrane transport, and protein synthesis in rat hepatocytes.

The cryopreservation of hepatocytes is of particular interest as a step in the possible treatment of some inborn disorders of metabolism. This study examines the metabolic damage that occurs as a result of the freeze-thaw procedures and during subsequent incubation periods of isolated rat hepatocytes. Even for freshly prepared hepatocytes, the presence of 1.8 M of Me2SO during incubation led to a rapid decline in viability. Optimal recovery after cryopreservation was obtained when incubation was started after the progressive removal of Me2SO. A buffer medium characterized by an intracellular electrolyte composition (Euro-Collins) proved particularly beneficial to the membrane integrity, probably by protecting the (Na+,K+)ATPase pump activity. The interpretation of viability using the trypan blue exclusion test was generally confirmed by the metabolic analysis of protein synthesizing activity and membrane transport function which are regarded as more rigorous tests of functional viability. The incorporation of L-[U-14C]isoleucine into the proteins of fresh hepatocytes during the first hour of incubation progressively leveled off over the next 2 hr. The cryopreserved hepatocytes showed a similar pattern although at a lower level of activity. Even after 3 hr of preincubation, the subsequent addition of labeled isoleucine still indicated a residual protein synthesizing activity. The active transport of alpha-amino[1-14C]isobutyric acid through the cell membranes reached a peak value after 60 min of incubation of fresh hepatocytes, and after 40 min of incubation of cryopreserved cells, followed by a steep decline as expression of rapid membrane deterioration. Again, the membrane transport pattern for the cryopreserved samples occurred at a lower level of activity. After preincubation of fresh and cryopreserved hepatocytes for 180 min, subsequent addition of labeled alpha-aminoisobutyric acid did not show any further significant metabolic activity. Initially the amino acid availability appeared to control protein synthesizing activity while, as membrane transport became seriously damaged, incorporation leveled off with only a low metabolic activity remaining. Although cryopreserved hepatocytes were susceptible to faster deterioration during subsequent incubation, considerable metabolic activity was retained. However, fresh and cryopreserved hepatocytes expressed metabolic functions at significantly different activities. Moreover, the differences between fresh and cryopreserved cells varied with the particular cellular function being examined.

Biochemical and functional aspects of recovery of mammalian systems from deep sub-zero temperatures.

The viability of isolated mammalian systems is, apart from possible morphological changes, essentially conditioned by the biochemical modifications from normal physiological conditions to an artificial environment where blood supply is interrupted leading to ischaemia and where the temperature is lowered. In order to survive freezing and thawing, mammalian systems have to be protected by cryoprotectants, which apart from some inherent toxicity, may also interact with vital metabolic mechanisms (Conover, 1969, 1975: Fahy, 1986: Fahy et al. 1984: Jacobs & Herschler, 1986: Karow, 1982: Penninckx et al. 198 3: Polge et al. 1949: Rowe et al. 1980: Schlafer, 1981: Taylor & Pignat, 1982). Cellular volume changes as a result of modifications in extra- a and intracellular osmolality occurring during freezing and thawing prove particularly detrimental to the normal functioning of the cellular membranes (Crowe et al. 1983: Farrant, 1980: Farrant et al. 1977b: Karow, 198 2: Mazur & Rigopoulos, 1983: Meryman, 1970: Meryman et al. 1977: Nei, 19 76: Santarius & Giersch, 1983). Furthermore intracellular ice formation enhances structural and metabolic injury to subcellular particles(Farrant et al. 1977a: Fink, 1986: Fishb ein & Griffin, 1976: Fujikawa, 1981: Fuller & De Loecker, 1985: Lazarus et al. 1982: Malinin, 1972: Mazur, 1984: Pavlock et al. 1984:Penninckx et al. 1984: Persidsky & Ellet, 1971: Rubinacci et al. 1986: Shikama, 1965: Steponkus & Wiest, 1979: Strauss & Ingenito, 1980: Takehara & Rowe, 1971: Tamiya et al. 1985). Even with the protection of structural integrity, the preservation of energy production and the maintenance of the specific intracellular medium are essential to secure viability (Pegg, 1981).