Presumptive congenital radial head sub-luxation in a shih tzu: successful management by radial head ostectomy.

Congenital radial head sub-luxation was diagnosed in a 7-month-old, neutered male shih tzu that presented with a limb deformity and severe lameness of the right fore limb. Radiography revealed a craniolateral sub-luxation of the right radial head, which was treated by radial head ostectomy, fixation of the radius to the ulna with a screw and joint stabilisation with suture-anchors and cerclage wire. Surgical treatment followed by physiotherapy resulted in a fully functional, well-aligned and non-painful elbow. To the authors' knowledge this is the first case report of a congenital radial head sub-luxation in a craniolateral direction in a dog and also one successfully managed with radial head ostectomy and radioulnar synostosis.

Canine distal hindlimb soft tissue sarcoma--a novel approach to surgical defect closure.

A 10-year-old, male, neutered whippet was presented with a soft tissue mass located on the medial aspect of the distal right tibia. The mass was 4 cm in diameter and of two months duration. Recent biopsy by the referring veterinarian, prompted by noticeable enlargement, identified the mass as a soft tissue sarcoma. Staging assessments did not reveal any evidence of metastases. Marginal excision was performed. The resultant defect was closed primarily by the creation of a bipedicle flap on the distal caudo-lateral aspect of the crus to enclose the Achilles tendon separately, leaving a defect between the Achilles tendon and the tibia. Postoperative management entailed support dressings and exercise restriction. Complete wound healing was attained three weeks postoperatively with excellent return of function. No recurrence was noted at eight months post-resection.

Canine meningioma: a case report of a rare subtype and novel atlanto basioccipital surgical approach.

OBJECTIVE: To describe a novel histological subtype of skull base and cranial cervical meningioma operated upon through an uncommon atlanto basioccipital approach. STUDY DESIGN: Clinical case report. ANIMAL: A seven-year-old neutered female Boxer. METHODS: The dog was presented due to lethargic behavior, signs of head and neck pain, and ongoing tetraparesis. Neurological examination and magnetic resonance imaging led to diagnosis of a ventral spinal canal mass causing severe cord and brainstem compression and which required surgical management. RESULTS: The mass was approached through a ventral craniectomy extending cranially on the basioccipital bone. Extemporaneous cytological examination of the mass led to confirmation of neoplastic nature of the abnormal tissue. Cytoreduction of the mass was performed using multiple cycles of curettage, lavage and suction. Histopathological examination of the tumour was consistent with a papillary variant of meningioma. Adjuvant therapy was declined by the owners. The dog improved considerably in the postoperative period, however three months after surgery signs of recurrence led to euthanasia. CONCLUSIONS: Basioccipital craniectomy allowed a limited but sufficient approach for cytoreduction of a compressive meningioma of the brainstem and cranial cervical spinal cord. Papillary meningiomas are locally infiltrative and postoperative recurrence is common. CLINICAL RELEVANCE: Basioccipital approach combined with cranial atlanto corpectomy can be performed to relieve compression of the caudal brainstem and cranial cervical cord.

Serum interleukin-6 and thyroid hormones in rheumatoid arthritis.

Using rheumatoid arthritis (RA) as a model, we have investigated whether the activation of the cytokine system, in particular, activation of interleukin (IL)-6 production, is a major cause of the depressed serum T(3) seen frequently in the nonthyroidal illness syndrome (NTIS). RA was chosen because it is a chronic autoimmune disease leading to increased serum IL-6 concentrations. We studied 16 untreated RA and 35 treated RA patients. Twenty-seven treated and 27 untreated patients with noninflammatory musculoskeletal symptoms served as controls. The patient groups displayed similar age distribution and nutritional status. Untreated RA patients displayed elevations of serum IL-6 (mean, 37.5 pg/mL) and C-reactive protein (CRP; mean, 41.3 mg/L), consistent with the inflammatory nature of their disease. Treated RA patients had significantly reduced serum IL-6 (mean, 9.9 pg/mL) and CRP (mean, 13.3 mg/L) compared with untreated RA patients, while untreated and treated patients with noninflammatory musculoskeletal symptoms had near normal serum IL-6 (mean, 2.5, 6.6 pg/mL, respectively) and CRP levels (mean, 5.8, 8.1 mg/L, respectively). However, there were no significant differences in serum concentrations of free T(3) (FT(3)) and free T(4) (FT(4)) between groups, and thyroid indices were in the normal range in RA patients. Moreover, no significant correlations between serum concentration of IL-6 and any of the thyroid hormones were demonstrated for any of the patient groups. In conclusion, we have been unable to confirm in RA that IL-6 activation leads to the low T(3) state of NTIS.

Endocrine and cytokine changes during elective surgery.

Elective surgery was used as a model of severe non-thyroidal illness (SNTI) to study the inter-relation between changes in serum thyroid hormones, thyroid stimulating hormone (TSH), cortisol, and interleukin 6 concentrations. The study was designed to determine whether the expected interleukin 6 increases after surgery are the cause of decreased serum tri-iodothyronine (T3) concentration normally observed following severe trauma. Blood was sampled for 24 hours before, during, and for 48 hours after abdominal surgery under general anaesthesia in 11 patients. Total T3 decreased 30 minutes after induction and continued to decrease at 24 hours. After a transient increase at 30 minutes, free T3 also decreased, and free thyroxine (T4) concentrations, other than a similar transient increase, did not change. TSH concentrations were increased at four hours and the nocturnal surge was suppressed. The increase in the serum interleukin 6 concentration was not observed until four hours. Cortisol concentrations were increased at 30 minutes and peaked at four hours. Therefore, the early changes in thyroid hormones and TSH accompanying surgery do not seem to be caused by changes in interleukin 6 concentrations.

Tumor necrosis factor-mediated cell lysis in vitro: relationship to cAMP accumulation and guanine nucleotide-binding proteins.

We have investigated the modulation of tumor necrosis factor (TNF)-mediated tumor cell lysis by cAMP. Among a panel of human breast tumor cell lines, MCF7 and MDA MB 231 were shown to be, respectively, sensitive and resistant to TNF-mediated cell lysis in vitro. 125I-labeled TNF-binding experiments demonstrated that both cell lines bind TNF, indicating that the differential sensitivity to TNF was not related to TNF receptor expression. To study the relationship between TNF-mediated cell lysis and cAMP accumulation, cAMP measurement was performed following TNF treatment. Our data show that TNF alone did not induce an enhancement of intracellular cAMP accumulation either in the TNF-sensitive or in the TNF-resistant cell line. Experiments in which cells were exposed to forskolin revealed that this cAMP elevating drug was efficient in enhancing the sensitivity to TNF of MCF7 cell line. This potentiating effect of forskolin was maximal for suboptimal concentrations of TNF (10 ng/ml), reaching up to 100% when forskolin was added at 100 microM. However, co-stimulating with forskolin of either MDA MB 231 or a TNF-resistant MCF7 clone (MCF7-R-A1) did not induce any reversal of resistance to TNF. We further assessed the interaction of TNF with transmembrane signalling and the possible involvement of guanine nucleotide-binding proteins (G-proteins). Bacterial toxin-catalyzed ADP ribosylation of MCF7 and MDA MB 231 membranes was, therefore, performed. Using cholera toxin, we demonstrate that TNF treatment did not quantitatively alter the activity of stimulatory G-proteins either in MCF7 or MDA MB 231 cell line. In contrast, pertussis toxin-catalyzed ADP ribosylation experiments suggest a functional coupling of TNF receptors to a 40-kDa pertussis toxin-sensitive G-protein in the TNF-sensitive MCF78 cell line but not in the TNF-resistant MDA MB 231 cell line. Taken together, these data indicate that cAMP might play a role in TNF-mediated cell lysis and are in support of the involvement of a pertussis toxin-sensitive G-protein in TNF-mediated MCF7 cells lysis.

Phorbol myristate acetate inhibits phosphoinositol lipid-specific phospholipase C activity via protein kinase C activation in conditions inducing differentiation in HL-60 cells.

We have studied, in streptolysin O-permeabilized HL-60 cells and in HL-60 membrane preparations, the effects of phorbol 12-myristate 13-acetate (PMA) on polyphosphoinositide-specific phospholipase C (PLC) activity and on terminal differentiation towards macrophagic-like cells. We showed that terminal differentiation was induced when differentiating concentrations of the drug were present for only 1-2 h in the culture medium. Conditions inducing differentiation also inhibited PLC activity for a long lasting period (at least 5 h). When terminal differentiation affected only part of the cell population, inhibition of phospholipase C activity was found to be less marked and reversible over the period studied. Moreover in experiments done in an HL-60 clone resistant to PMA, no inhibition of PLC activity was provoked by this tumour promotor. In order to study the involvement of protein kinase C in this process, we measured modifications of PLC activity by PMA in the presence of two different protein kinase C inhibitors, staurosporine and H-7. They both prevented the inhibition of PLC activity by PMA indicating that this inhibition is likely to be related to the effect of PMA on protein kinase C activity. This was also confirmed by the fact that active protein kinase C, by itself, was able to decrease PLC activity when added to membrane preparations or to streptolysin O-permeabilized control HL-60 cells. These results indicate that PMA acts in inhibiting phospholipase C activity through its effect on protein kinase C activation and/or on protein kinase C translocation to the plasma membrane and that terminal differentiation, might be related to changes in both protein kinase C and PLC activities.

The differentiating agent, retinoic acid, causes an early inhibition of inositol lipid-specific phospholipase C activity in HL-60 cells.

Retinoic acid, a derivative of vitamin A, is shown to inhibit the levels of inositol phosphates and diacylglycerol by 25-30% when added to intact HL-60 cells at concentrations which induce differentiation. The onset of inhibition occurs after 10 min and reaches a maximum at 45 min. To study the mechanism and the site of action of retinoic acid, the activity of the phosphatidylinositol bisphosphate-specific phospholipase C was studied in cells permeabilized with streptolysin O and in membrane preparations. Phospholipase C activity was stimulated either via the guanine nucleotide regulatory protein (G-protein) or directly by Ca2+. Retinoic acid treatment, in a time- and concentration-dependent manner, led to a decrease in phospholipase C activity when stimulated with either GTP gamma S or NaF, both of which activate the enzyme via the G-protein. By contrast, it had no effect on the enzyme activity when stimulated with Ca2+ alone. This indicates that retinoic acid interferes with the coupling of the G-protein and phospholipase C. A relationship between the inhibition of phospholipase C activity and the induction of differentiation by retinoic acid was investigated. Only a small inhibition of GTP gamma S-stimulated phospholipase C activity was observed when an analogue of retinoic acid, etretine or Ro10-1670, with low differentiating activity, was used. Moreover, no inhibition of the GTP gamma S-stimulated phospholipase C activity was observed in an HL-60 sub-line resistant to retinoic acid. These results suggest that phospholipase C inhibition is an important step in the induction of differentiation.

Use of liquid culture and cell cycle analysis to compare drug damage following in vitro treatment of normal human bone marrow cells with adriamycin, arabinosyl-cytosine, and etoposide.

The effects of adriamycin (ADM), arabinosyl-cytosine (ARA-C), and etoposide (VP16) were studied on human bone marrow mononucleated cells using colony formation in agar, a modified liquid culture system, and flow cytometry analysis of the cell cycle. Drug concentrations tested during a 1-h incubation ranged from 0.1 to 4 micrograms/ml for ADM, from 0.3 to 30 micrograms/ml for VP16, and from 10(-7) to 10(-3) M for ARA-C. Regression analysis of the dose-response curves was used to assess the drug concentration that inhibited 90% +/- 5% (LD90) of colony growth. LD90s were 0.4 microgram/ml for ADM, 20 micrograms/ml for VP16, and 10(-4) M for ARA-C. LD90-surviving cells were cultured in liquid medium for 3 weeks. Surviving cells over this time were 13% of the control for ADM, 22% for VP16, and 95.7% for ARA-C. Although cells decreased drastically in ADM- and VP16-treated samples, granulocyte-macrophage colony-forming units (CFU-GM) per 10(5) surviving cells rose to twice the control for ADM, to 60% for VP16, and to 150% for ARA-C. Flow cytometry analysis of the cell cycle was performed at day 0 and at day 4 after treatment with the LD90 dose. It showed a rapid and reversible effect of ARA-C on cells in the S-phase, whereas the action of VP16 concerned all cells, regardless of their cycle phase. We conclude that the direct effects of the three drugs on CFU-GM in agar are poorly predictive of hematopoietic reconstitution capacity, except for VP16. Liquid culture gives a much more accurate appraisal of the long-term damage and recovery due to anticancer drugs.

Effect of hematocrit concentration on blood glucose value determined on Glucometer II.

Nineteen patient blood samples each with modified hematocrit concentrations of approximately 20, 30, 40, 50, and 60%, were assayed for their glucose concentration by the Glucometer II. Blood removal from the test strip was by the one- and two-blot techniques. The reference method was the Yellow Springs Instruments (YSI) blood glucose analyzer. Glucometer II results were falsely high for the single blot (13-59%, mean 33%) and double blot (12-41%, mean 19%) at 20% hematocrit and falsely low at 60% hematocrit for the single blot (22-44%, mean 37%) and the double blot (26-49%, mean 43%). At 40-50% hematocrit, Glucometer II and YSI results agreed only for the one-blot technique.

The effect of the haematocrit value on the determination of glucose levels by reagent-strip methods.

Twenty-one patients' blood samples, each with modified haematocrit concentrations of about 20%, 30%, 40%, 50% and 60%, were assayed for the presence of glucose by two reagent-strip methods--the Ames Glucometer with BG reagent strips and the Reflolux reflectance meter with BM-test strips. The reference method was the YSI blood glucose analyser. Both Ames and BM systems were found to have good precision. Ames results were falsely high (mean, 15%) at a haematocrit value of 20% and falsely low (mean, 21%) at a haematocrit value of 60%. BM results also overestimated (mean, 4%) and underestimated (mean, 5%) at the corresponding haematocrit levels. The results showed good agreement between Ames and BM methods and the YSI method for normal haematocrit concentrations of 40% to 50%. Clinicians should be aware of inaccuracies in glucose results by test-strip methods in patients with abnormal haematocrit concentrations.

Differentiation of mixed colony-forming cells in normal human bone marrow and blood.

In blood and marrow samples from ten normal adults, mixed colonies were found in agar cultures at concentrations of 4.1 +/- 3.4/10(5) mononuclear bone marrow cells, and of 4.7 +/- 4/5 X 10(5) mononuclear blood cells. Reseeding of five- to seven-day clusters of immature cells in fresh medium with 5% PHA leukocyte-conditioned medium (PHA-LCM) and 2 U erythropoietin (Epo) showed that 10(5) mononuclear cells contained as many as 42 +/- 12 CFU-mix in bone marrow and 12 +/- 7 CFU-mix in blood, i.e., ten times more than in the primary cultures. Attempts were made to discover why these potentially mixed colonies failed to develop in primary cultures. For this purpose five- to seven-day clusters of immature cells from cultures of 5 X 10(4) bone marrow cells grown with PHA-LCM, Epo, or both were reseeded in medium containing one or both stimulants. The presence of both in secondary culture gave the best CFU-mix growth, whatever the primary stimulation, i.e., more than one-third of the immature clusters gave rise to colonies of red cells and granulocyte lineages. The addition of one or both stimulants between days 7 and 14 of primary culture, however, did not increase the number of mixed colonies or give rise to late developing ones. Reseeding of colonies and clusters of 14-day primary cultures showed that at least one-third of erythroid bursts or of late-developing granulocyte colonies contained CFU-mix, but that less than 10% of 14-day clusters developed into mixed colonies. Cultures of nonadherent cells, mononuclear cells of less than 1.065/g density or with a velocity sedimentation rate of less than 6 mm/h gave the same results as the culture of total mononuclear cells; i.e., at least ten times more multipotent progenitors were found by reseeding after five to seven days than in primary cultures. We conclude that in human bone marrow and blood the physical and kinetic properties of pluripotent stem cells are similar to those of 14-day erythroid bursts and GM colonies. Under normal culture conditions, most of these cell types differentiate into one cell lineage only, and plurilineage differentiation is inhibited.