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Pressure-induced ischemia. I. An experimental model for intravital microscopic studies in hamster cheek pouch.

A model to study pressure-induced ischemia by intravital microscopy is presented. A hamster cheek pouch is prepared to get a single layer of epithelium, together with vessels and connective tissue. Pressure, which can be varied, is transmitted to the tissue by a pressure chamber with a rubber membrane. Microcirculatory reactions may be studied while the pressure is applied, when the pressure is released and as the tissue is regaining circulation. The local environment is controlled by irrigating the test tissue with a solution approximating the composition of interstitial fluid. Local oxygen and carbon dioxide tensions are controlled. There is a marked difference in restoration of blood flow to the tissue after 2- and 4-hour ischemia. After 2-hour ischemia the tissue regained circulation rapidly. Microbleedings developed during the postpressure observation time. After 4-hour ischemia, on the other hand, the tissue regained circulation slowly and only in one third of the microvessels. Extensive white blood cells sticking to the vessel walls were seen indicating endothelial damage. In the 4-hour experiments there were very few microbleedings compared to the 2-hour experiments.

Animals↗

The cheek teeth of the rabbit: morphology, histology and development.

A detailed anatomic and histologic description of the cheek teeth of the rabbit, both deciduous and permanent, is presented, together with the chronology of their pre- and postnatal development. The deciduous teeth are of limited growth, while the permanent ones grow continuously. Certain aspects of cementogenesis in these teeth are discussed.

Animals↗

Effects of buflomedil on spontaneous vasomotion and mean arteriolar internal diameter in the hamster cheek pouch.

Intravital microscopy of the hamster cheek pouch microvasculature was used for in vivo studies on the effects of buflomedil, phentolamine (alpha-adrenergic receptor antagonist) and norepinephrine on the mean internal arteriolar diameter and spontaneous vasomotion. All drugs were applied topically. The vasomotor activity was studied in 125 arterioles (internal diameter range 18.0-62.0 microns) of 34 preparations. Addition of buflomedil (10(-9) to 10(-5) M) did not affect the arteriolar diameter significantly (from 100.7 +/- 3.5 to 106.4 +/- 1.8%, values expressed in percent of the initial diameter as mean +/- SE), but increased the vasomotion frequency and amplitude by approximately 20% (from 7.5 +/- 0.3 to 9.2 +/- 0.2 cpm) and 30% (from 7.3 +/- 0.3 to 10.0 +/- 0.5 micron), respectively. Phentolamine (10(-9) to 10(-5) M) dose-dependently increased the microvascular diameter (from 102.3 +/- 1.2 to 139.1 +/- 4.3%) and reduced the vasomotion frequency and amplitude (from 8.0 +/- 0.3 to 1.9 +/- 0.5 cpm and from 9.0 +/- 2.1 to 3.1 +/- 0.2 microns, respectively). Addition of buflomedil (10(-7) M) reduced the vasodilation evoked by phentolamine (from 103.3 +/- 0.7 to 127.0 +/- 1.5%) and potentiated its depressive effect on vasomotion frequency and amplitude (from 7.6 +/- 0.1 to 1.0 +/- 0.3 cpm and from 9.0 +/- 0.3 to 1.9 +/- 0.6 microns, respectively). Norepinephrine (10(-9) to 10(-5) M) dose-dependently decreased to arteriolar diameter (from 102.3 +/- 0.7 to 69.6 +/- 1.6%) and the vasomotion frequency and amplitude (from 8.4 +/- 0.2 to 0.4 +/- 0.3 cmp and from 8.7 +/- 0.2 to 0.5 +/- 0.4 microns, respectively).(ABSTRACT TRUNCATED AT 250 WORDS)

Administration, Topical↗

Vasoactive effects of basic and acidic fibroblast growth factors in hamster cheek pouch arterioles.

Fibroblasts growth factors (FGFs) exhibit well-known angiogenic actions, but there is some controversy about whether they have vasoactive effects on blood vessels which might contribute to angiogenesis per se. To clarify this, changes in arteriolar diameter were recorded during observation by videomicroscopy of 3rd- and 4th (terminal)-order arterioles (resting diameters 22.5 +/- 0.5 microns and 14.4 +/- 0.3 microns, respectively) in the hamster cheek pouch in response to FGF application. Recombinant human bFGF (basic) and aFGF (acidic) were applied from micropipettes positioned 5-10 microns from the adventitial surface of vessels. Maximum vasodilator effects of adenosine (10(-4) M) applied in a similar way were also observed. Adenosine increased the diameters of 4th-order arterioles by 37.2 +/- 3.8% and those of 3rd-order arterioles by 38.7 +/- 2.7. bFGF produced vasodilatation (threshold dose 0.1 ng ml-1) in both classes of arterioles, while aFGF produced dose-dependent constriction (threshold dose 0.01 ng ml-1). A maximal dilator effect in 4th-order arterioles was obtained with 100 ng ml-1 bFGF, when diameters reached 82.6 +/- 2.4% of those with adenosine. Maximal constrictor effect (-48.2 +/- 5.6% of resting diameter) occurred with a dose of 100 ng ml-1 aFGF. Vehicle alone (MOPS or bicarbonate buffer used as solvents for FGFs) had no effect. As vasoconstrictors are known to stimulate growth of smooth muscle cells while dilators stimulate growth of endothelial cells, it is possible that the opposing vasoactivities demonstrated for aFGF and bFGF are linked with their selective mitogenicity for smooth muscle and endothelial cells, respectively, and contribute to their angiogenic actions.

Adenosine↗

Correlation between laser Doppler perfusion monitoring and hematocrit in hamster cheek pouch microcirculation.

The aim of this study was to investigate the relationships between laser Doppler perfusion monitoring (LDPM) measurements and different systemic hematocrits in microcirculation in terms of changes in oscillatory flow patterns. The hamster cheek pouch microvasculature was visualized by a fluorescent microscopy technique, and LDPM signals were derived from arterioles and venules under control conditions and after isovolemic hemodilution with saline and 6% dextran, MW 70,000 to 26.1 +/- 2.1%. Vasomotion, oscillations of microvascular blood flow (flow motion) and red blood cell (RBC) velocity were analyzed with Fourier transform and autoregressive modeling. LDPM recordings presented a significant increase in perfusion units (PU) during hemodilution-184 +/- 15 versus baseline 137 +/- 11 PU in arterioles and 40.2 +/- 3.5 versus 28.6 +/- 4.3 PU in venules-that was correlated with a significant increment in arteriolar and venular RBC velocity. There was a rise in the frequency [2.9 +/- 0.5 cycles per min (cpm) vs. 1.8 +/- 0.5 cpm] and spectral power of flow motion in arterioles whereas the increase in spectral power was related to a decrease in frequency (12.6 +/- 2.1 vs. 3.6 +/- 0.7 cpm) in venules. Oscillations in arteriolar and venular RBC velocity revealed coincident frequency components with flow motion patterns. The present data suggest that the LDPM measurements are more sensitive to velocity than hematocrit. Furthermore, hemodilution appears to affect differently arteriolar and venular flow motion patterns.

Animals↗

Platelet aggregation in arterioles of the hamster cheek pouch and in heart transplants: its tissue-dependent influencibility by acetylsalicylic acid and nafazatrom.

Different, tissue-dependent antiaggregatory properties of endothelial cells are suggested by in vitro findings, which show different spectra and amounts of arachidonic acid products to be synthesized by vessels of different type and origin. Platelet aggregation was assessed in vivo in native arterioles of the hamster cheek pouch and in arterioles of heart transplants by intravascular excitation of fluorescein isothiocyanate-dextran (FITC-d). The time to aggregate appearance (TAA) was determined, i.e. the interval from onset of FITC-d excitation until the first aggregate was seen to adhere to the vessel wall. Two groups were pretreated with 100 micrograms/kg nafazatrom and 100 mg/kg acetylsalicylic acid (ASA), both potent antithrombotic drugs. Nafazatrom has been shown to stimulate prostacyclin release from endothelial cells, whereas ASA blocks cyclooxygenase both in platelets and vessel walls. TAA was the same for native arterioles of both types. Nafazatrom was equally effective in both vessels, and about three orders of magnitude more potent than ASA in prolonging TAA. ASA was twice as effective in arterioles of the heart as it was in those of the skin type, indicating that tissue-dependent mechanisms achieve the antiaggregatory potency of the vessel walls. It is concluded that further in vivo studies on platelet aggregation and thrombus formation should be performed on cardiac vessels directly and not on vessels of different origin. The presented model may be a useful tool for investigating the mentioned tissue-dependent mechanisms of thrombus formation in vivo.

Animals↗

Homocellular conduction along endothelium and smooth muscle of arterioles in hamster cheek pouch: unmasking an NO wave.

We investigated roles for homocellular (endothelium or smooth muscle) and heterocellular (myoendothelial) conduction pathways along hamster cheek pouch arterioles in vivo (n=64; diameter, 33+/-1 microm). Endothelium-dependent and -independent vasoactive agents were delivered from micropipettes (0.5 or 1 second pulse) onto an arteriole while observing diameter changes at defined distances along the vessel. Acetylcholine (ACh) produced maximal diameter (63+/-1 microm) locally and vasodilation conducted rapidly ( approximately 10 microm response at 2 mm, <1 second). Responses to bradykinin (BK) were similar, whereas sodium nitroprusside produced maximal dilation locally without conduction. KCl evoked biphasic conduction of vasoconstriction and vasodilation, whereas phenylephrine (PE) produced conducted vasoconstriction. Disrupting the integrity of endothelium as a conduction pathway using focal light-dye treatment (LDT) abolished conducted vasodilation to BK and to KCl but not to ACh. Disruption of smooth muscle integrity with LDT abolished conducted vasoconstriction with no effect on conducted vasodilation. After LDT of respective cell layers at sites 1 mm apart, vasodilation to ACh conducted past disrupted smooth muscle or disrupted endothelium, but not beyond both sites in series. The loss of conduction after selective LDT indicates a lack of effective myoendothelial coupling along the arteriolar wall. During NO synthase inhibition (L-NA, 100 micromol/L), conducted vasodilation was abolished to BK and to KCl yet remained intact to ACh. However, after LDT of smooth muscle, L-NA inhibited conduction to ACh by 60%. Thus, conduction of vasodilation entails a wave of NO release along arteriolar endothelium that is masked when smooth muscle provides a parallel conduction pathway.

Acetylcholine↗

Vasoconstrictor effects of platelet-activating factor in the hamster cheek pouch microcirculation: dose-related relations and pathways of action.

Platelet-activating factor (PAF) has been implicated as a potential mediator of inflammatory processes. In this study, we quantified the effects of PAF on vessel diameter in a microvascular bed and investigated the biochemical pathways of this compound. The hamster cheek pouch microcirculation was observed with intravital microscopy. Experiments were video-recorded and analyzed with an image shearing device. Vasoconstriction was the predominant vasomotor response to PAF. PAF (10(-10) -10(-5) M) was applied topically to the pouch for 3 minutes. Arterioles ranging in size from 8 to 15 micron were the most sensitive, and they constricted completely in response to PAF 10(-7) and 10(-5) M. Arterioles 21-40 micron in diameter constricted to 12-17% of control after PAF at 10(-7) and 10(-5) M, respectively; they reopened to about 70% of their control value after a few minutes and remained near that size throughout the experiment. Arterioles 41-60 micron in diameter constricted to about 20% control size in response to 10(-7) and 10(-5) M PAF, and by the end of the experiment, these vessels had returned to about 90% control size. To determine the pathways of PAF actions, inhibitors of the arachidonic acid cascade and receptor blockers were used. Dexamethasone, indomethacin, OKY-046 (a thromboxane A2 synthetase inhibitor), and kadsurenone (a PAF-receptor blocker) blocked the vasoconstrictor response to PAF. Our experiments demonstrate that PAF-produced arteriolar constriction in a microvascular bed is 1) dose-related, 2) dependent upon vessel size, 3) largely due to thromboxane A2 activity, and 4) mediated by PAF-receptor interactions.

Animals↗

Effect of platelet-activating factor on microvascular permselectivity: dose-response relations and pathways of action in the hamster cheek pouch microcirculation.

Platelet-activating factor (PAF) has recently been described as a mediator of inflammatory processes. In this study, we quantitated the dose-response effects of topically applied PAF on microvascular permselectivity and investigated the biochemical pathways of this compound. Permselectivity alterations were assessed by measuring the clearance of macromolecules with fluorescein isothiocyanate dextran 150 (FITC-dx 150) as a tracer. The microvascular bed of the hamster cheek pouch served as a model. PAF was found to induce leakage of macromolecules from postcapillary venules. Control FITC-dx 150 plasma clearance (+/- SEM) was 72 +/- 10 nl/90 min. Clearances of 61 +/- 10 474 +/- 145, 622 +/- 57, 301 +/- 86, and 142 +/- 3 nl/90 min were obtained at PAF concentrations of 10(-9), 10(-8), 10(-7), 10(-6), and 10(-5) M, respectively. A one-way analysis of variance showed that the population means were not equal. Multiple comparison by the Student-Newman-Keuls test demonstrated that the clearances obtained with 10(-8), 10(-7), and 10(-6) M were significantly greater than controls. Significant differences existed between 10(-7) M PAF and 10(-9), 10(-6), and 10(-5) M PAF. In an effort to elucidate the biochemical pathways of PAF activity, several inhibitors of the arachidonic acid cascade and receptor blockers were used. Dexamethasone and kadsurenone attenuated the clearance response to PAF in a statistically significant manner, while indomethacin, OKY-046, and chlorpheniramine were without effect.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗

Chemodectoma of the cheek. A case report.

The first case of a chemodectoma of the cheek is presented and the histopathology reviewed. Chemodectomas of the head and neck can be difficult to diagnose, with angiography remaining the single most useful aid to determining both location and diagnosis. Surgery is the treatment of choice, and prognosis depends not only on the extent of the tumor but also its predilection for malignant behavior.

Adult↗

Immunotherapy of chemically-induced tumors in the hamster cheek pouch with dinitrochlorobenzene.

Immune stimulation with an agent such as dinitrochlorobenzene (DNCB) may delay chemical carcinogenesis. Dimethylbenzanthracene (DMBA) was used to chemically induce tumors in the hamster buccal pouch. Hamsters were studied for the effect of DNCB sensitization in the buccal pouch prior to or after DMBA tumor induction. At appropriate time intervals the hamsters were sacrificed and each cheek pouch was examined histologically for the development of DMBA-induced tumors and for the presence of lymphoid cells infiltrating the tumor site. The results show that DNCB immunotherapy or immunoprophylaxis prior to or following DMBA tumor induction can alter the type of tumor produced and stimulate an infiltration of lymphoid cells into the tumor area probably invoking immune defense mechanisms.

9,10-Dimethyl-1,2-benzanthracene↗

The effect of vitamin A on hamster cheek pouch mucosa in organ culture.

Vitamin A was added to the medium of neonatal hamster cheek pouch mucosa maintained in organ culture. The epithelium of the treated explants showed a reduction in keratinization and down-growth into the underlying connective tissue. The cytological and morphological changes in the epithelium were consistent with glandular metaplasia.

Animals↗

Changes in the microvasculature after mechanical pressure on the hamster cheek pouch.

The reaction of the microvasculature in the periodontal ligament to mechanical pressure is considered a very important phenomenon with respect to the biological background to orthodontic tooth movement. For clarification of the microvascular reaction to mechanical pressure, an experimental model that incorporated a hamster cheek pouch was established. This in vivo model solves some of the problems of other experimental models of the microvasculature of the periodontal ligament. Blood plasma permeation in this model was examined by means of a blue dye (pontamine sky blue), and histological observation was performed by light and transmission electron microscopy. Increased vascular permeability was observed within 30 min after removal of the mechanical stimulation. The threshold weight resulting in dye leakage was between 1 and 5 g applied for 60 min. From the histological observations, some large gaps between the endothelial cells in venules were found, while most basement membranes remained undisturbed. These observations suggest that the plasma leakage was due to mild traumatic injury to the endothelial cells. Both leukocytes and platelets were observed in the gaps between the endothelial cells in the venule. These types of microvascular reactions to mechanical pressure could initiate tissue changes in the periodontal ligament during orthodontic tooth movement.

Animals↗

Differential effects of noxious conditioning stimulation of the cheek by capsaicin on human sensory and inhibitory masseter reflex responses evoked by tooth pulp stimulation.

In this study, we investigated whether selective activation of nociceptive primary afferent fibers by capsaicin would induce modulations on tooth-pulp-evoked sensory or inhibitory masseter reflex responses in healthy human subjects. The contribution of central N-methyl-D-aspartate (NMDA) receptor mechanisms in capsaicin-induced effects on sensory or reflex responses was evaluated by dextromethorphan, an NMDA-receptor antagonist. The inhibitory masseter reflex was evoked by electrical stimulation (constant current, single pulses) of the upper incisor while the subject was biting at 10% of his maximal force. The sensation of the tooth pulp stimulation was evaluated by visual analogue scale (VAS). The magnitude, duration, and the the latency of the reflex were determined by bite force measurements. The inhibitor masseter reflex could be induced by non-painful tooth pulp stimulation, and the inhibition was enhanced as a function of increasing stimulus intensity. Capsaicin (1%) applied topically to the skin of the cheek produced a spontaneous burning pain sensation. During capsaicin treatment, the VAS ratings for the sensation induced by tooth pulp stimulation were significantly reduced, whereas no significant changes were found in the tooth-pulp-induced masseter reflex responses. Double-blind treatment with dextromethorphan at a dose of 100 mg (= the highest does without side-effects) had no effect on sensory or reflex responses. These data indicate that noxious stimulation of the facial skin by capsaicin induces differential effects on tooth-pulp-evoked sensory and inhibitory masseter reflex responses: Sensory responses are strongly attenuated, while masseter reflex responses are not significantly changed. Dextromethorphan at a clinically applicable dose does not influence tooth-pulp-evoked sensory or reflex responses or their modulation by capsaicin. Furthermore, the lack of modulation of the masseter reflex response by capsaicin differs from the capsaicin-induced enhancement of a nocifensive limb flexion reflex described earlier.

Administration, Cutaneous↗

The combined masseter muscle/intraoral cheek transposition (IOCT) flap for primary reconstruction of the dorsal oral cavity.

A method for primary reconstruction of the dorsal oral cavity after tumor resection of T2 or small T3 tumors is presented. By combination of the masseter muscle flap with a cranial pedicled intraoral cheek transposition (IOCT) flap, a reconstruction of the defects with two layers of tissue is possible. This easy and functional reconstruction method is especially useful for high-risk patients because of the markedly reduced operating time.

Aged↗

Vasoconstrictive effects of cocaine and lidocaine with epinephrine in the hamster cheek pouch model.

Otolaryngologists frequently use cocaine and lidocaine with epinephrine for their anesthetic and vasoconstrictive effects. However, no studies have visually compared the vasoconstrictive effects of cocaine to lidocaine with epinephrine. This study used the everted hamster cheek pouch model to assess the vasoconstrictive effects of cocaine and lidocaine with epinephrine by measuring the diameter changes of arterioles. Our results showed that locally injected 1% lidocaine with 1:100,000 units epinephrine caused the greatest diameter reduction in arterioles to 32% of the initial resting diameter. Topical 1% lidocaine with 1:100,000 units epinephrine caused a gradual reduction in diameter with findings equivalent to those of injected 1% lidocaine with 1:100,000 units epinephrine at 30 minutes. Topical 5% cocaine HCl showed a minimal response, with a reduction to only 85% of the resting arteriolar diameter. Topical NaCl and injected NaCl controls showed no significant arteriolar reduction. Vasoconstrictive differences between the preparations were statistically significant (p < 0.05). Five percent cocaine is a much weaker vasoconstrictor than locally injected 1% lidocaine with 1:100,000 units epinephrine.

Administration, Cutaneous↗

Desmosomes in hamster cheek pouch epithelium: their quantitative characterization during epithelial differentiation.

Desmosomes in stratified squamous epithelia appear to exhibit quantitative alterations during differentiation. In this work we use stereological and other morphometric methods to quantify these structures in epithelial cells from defined basal, spinous and granular strata. Hamster cheek pouch mucosa from five animals was processed for electron microscopy using strictly standardized techniques and a stratified random sampling procedure was used to obtain micrographs of cells from basal, spinous and granular layers. Stereological intersection counting techniques were used to determine for each layer the relative surface area of plasma membrane occupied by desmosomes (Ss), the number of desmosomes per unit surface area of plasma membrane (Ns), the mean individual desmosomal diameter (delta) and the mean individual desmosomal surface area (s). In addition, estimates of nuclear volume were obtained by direct measurement of nuclear profiles and volume-to-surface ratios were obtained by a combination of point and intersection counting, which enabled estimates for the volume (Vcell) and plasma membrane surface area (SPM) of the 'average' cell within each stratum to be acquired. Using this information, it was then possible to calculate both the total surface area (S) and the number (N) of desmosomes on the plasma membranes of average cells. The parameters Ss and Ns showed progressive increases between basal and granular layers, whereas values for delta and s were lower in granular cells when compared with basal and spinous cells. The parameters Vcell, SPM, S and N all increased progressively and significantly during differentiation. Between basal and granular layers, the mean cell volume and surface area had each increased approximately threefold, whereas the surface area and number of desmosomes on the average cell plasma membrane had increased approximately seven- and eleven-fold, respectively. Granular cells thus possess more numerous desmosomes, which occupy a greater proportion of the plasma membrane area but which are individually smaller, when compared with basal and spinous layers.

Animals↗