Dissector Technique Research Articles

A new device to treat ping-pong skull fractures: The hammer puller technique. A comparative analysis using a realistic simulation model.

This study aims to describe a new surgical technique for the treatment of ping-pong skull fractures and to evaluate its efficacy in a realistic simulation model compared to the dissector elevation technique. A total of 64 fractures were obtained using 16 model units, each with four fractures (two frontal and two parietal). The hammer puller technique was applied for left-sided fractures and the dissector technique for right-sided fractures. The variables evaluated were fracture repair time, fracture volume, fracture corrected volume, and fracture correction percentage. Fractures were separated into groups according to the surgical technique used (hammer or dissector) and the bone fractured (frontal or parietal). Statistical analysis was performed with Jamovi® software (version 2.3) using Student's t-test. A complete degree of fracture correction was achieved with both techniques, demonstrating a sufficient performance in the correction of the deformity. The hammer technique was shown to be faster in correcting frontal bone depressions with 20.1 ± 7.8 s compared to 31.3 ± 4.7 s for the dissector technique, P < 0.001. There was no statistically significant difference for parietal applications (P = 0.405). This study describes a new minimally invasive surgical technique for the treatment of ping-pong fractures. Comparative analysis showed that both techniques were equally effective but that the hammer puller technique was more efficient than the dissector elevation technique, especially for frontal bone fractures.

Accurate Follicle Enumeration in Adult Mouse Ovaries

Sexually reproducing female mammals are born with their entire lifetime supply of oocytes. Immature, quiescent oocytes are found within primordial follicles, the storage unit of the female germline. They are non-renewable, thus their number at birth and subsequent rate of loss largely dictates the female fertile lifespan. Accurate quantification of primordial follicle numbers in women and animals is essential for determining the impact of medicines and toxicants on the ovarian reserve. It is also necessary for evaluating the need for, and success of, existing and emerging fertility preservation techniques. Currently, no methods exist to accurately measure the number of primordial follicles comprising the ovarian reserve in women. Furthermore, obtaining ovarian tissue from large animals or endangered species for experimentation is often not feasible. Thus, mice have become an essential model for such studies, and the ability to evaluate primordial follicle numbers in whole mouse ovaries is a critical tool. However, reports of absolute follicle numbers in mouse ovaries in the literature are highly variable, making it difficult to compare and/or replicate data. This is due to a number of factors including strain, age, treatment groups, as well as technical differences in the methods of counting employed. In this article, we provide a step-by-step instructional guide for preparing histological sections and counting primordial follicles in mouse ovaries using two different methods: [1] stereology, which relies on the fractionator/optical dissector technique; and [2] the direct count technique. Some of the key advantages and drawbacks of each method will be highlighted so that reproducibility can be improved in the field and to enable researchers to select the most appropriate method for their studies.

Comparison of methods for quantifying primordial follicles in the mouse ovary

BackgroundAccurate evaluation of primordial follicle numbers in mouse ovaries is an essential endpoint for studies investigating how endogenous and exogenous insults, such as maternal aging and chemotherapy, impact the ovarian reserve. In this study, we compared and contrasted two methods for counting healthy primordial follicles following exposure to cyclophosphamide (75 mg/kg), a well-established model of follicle depletion. The first was the fractionator/optical dissector technique, an unbiased, assumption-free stereological approach for quantification of primordial follicle numbers. While accurate, highly reproducible and sensitive, this method relies on specialist microscopy equipment and software, requires specific fixation, embedding and sectioning parameters to be followed, and is largely a manual process that is tedious and time-consuming. The second method was the more widely used serial section and direct count approach, which is relatively quick and easy. We also compared the impacts of different fixatives, embedding material and section thickness on the overall results for each method.ResultsDirect counts resulted in primordial follicle numbers that were significantly lower than those obtained by stereology, irrespective of fixation and embedding material. When applied to formalin fixed tissue, the direct count method did not detect differences in follicle numbers between saline and cyclophosphamide treated groups to the same degree of sensitivity as the gold standard stereology method (referred to as the Reference standard). However, when Bouin’s fixative was used, direct counts and stereology were comparable in their ability to detect follicle depletion caused by cyclophosphamide.ConclusionsThis work indicates that the direct count method can produce similar results to stereology when Bouin’s fixative is used instead of formalin. The findings presented here will assist others to select the most appropriate experimental approach for accurate follicle enumeration, depending on whether the primary objective of the study is to determine absolute primordial follicle numbers or relative differences between groups.

Volumetric investigation of the hippocampus in rat offspring due to diabetes in pregnancy–A stereological study

BackgroundThe brain development during the prenatal period is affected by various factors, including the mother's metabolic condition. It has been revealed that diabetes in pregnancy is associated with structural and functional alterations in offspring’s hippocampus. Hippocampus, as a critical region with well-known roles in learning and memory consolidation, is vulnerable to changes in glucose level. This study was designed to investigate the effects of maternal diabetes during the pregnancy period and insulin therapy on the neuronal density and the volume of different subfields of the hippocampus in rat offspring at postnatal day 14 (P14). MethodsWistar female rats were randomly divided into diabetics (STZ-D), diabetes treated with insulin (STZ-INS) group, and controls (CON). The animals in all groups were mated by non-diabetic male rats. Two weeks after birth, male pups from each group were sacrificed. The Cavalieri method was carried out to estimate the total volume, and the numerical density of the neurons in the hippocampus and its sub regions was measured by the optical dissector technique. ResultsBilateral hippocampal volume decreased in the diabetic group, mainly in the CA1, dentate gyrus (DG) and subiculum areas (P ≤ 0.05), when compared to control and insulin-treated diabetic animals. In all hippocampus sub-regions, maternal diabetes resulted in a significant decrease in the number of cells in comparison with two other groups (P ≤ 0.05 each). ConclusionThese data indicate that diabetes during pregnancy has a negative impact on the development of the hippocampus in the rats. These changes in the volume of hippocampal CA1, DG, and subiculum areas might be at the core of underlying neurocognitive and neurobehavioral impairments observed in the children of diabetic mothers.

ULTRASONIC DISSECTION VERSUS CONVENTIONAL LIGATION COAGULATION IN THYROIDECTOMY

ULTRASONIC DISSECTION VERSUS CONVENTIONAL LIGATION COAGULATION IN THYROIDECTOMY Abdulameer Muhsin Aldaraji MB, ChB, FICMS, General Surgeon, Al-Faiha General Hospital, Basrah, IRAQ. Abstract Ligation and diathermy coagulation was the standard method of hemostasis in thyroid surgery for more than a century. New hemostatic techniques were developed in the 1990th. Ultrasonic dissector used for the first time in thyroidectomy in 1999 by Tokami. Many studies compared the conventional method of thyroidectomy with new ultrasonic dissector technique. These studies found many advantages of the new method including reduction of operative time, shorter incision, decrease blood loss, decrease post operative drainage, shorten hospital stay and decrease complications. This study compared the two methods of hemostasis regarding incision length, operative time and complications. Between March 2012 and March 2015, 143 patients underwent open thyroidectomy involved in a prospective study at Al-Faiha General Hospital. The patients were divided into 2 groups: Group 1, conventional thyroidectomy technique(clamp, tie and diathermy coagulation) included 104 patients. Group 2, ultrasonic dissection technique (sutureless thyroidectomy) included 39 patients. The patient characteristic and postoperative complications were reported. The incision length and the operative time was measured. All patients underwent surgery by the routine pre tracheal strap muscle cutting transeversly (not splitting). In group 2, the ultrasonic dissector was used in strap muscle cutting, sealing §ion of the blood vessels and in the final resection of the gland. The surgical incision length and operative time were compared in both groups. In lobectomy, there was no significant reduction in the length of incision with use of ultrasonic dissector( z value0.05), while in subtotal, total or near total thyroidectomy there was significant reduction in the length of incision (p value

Lifestyle Factors Associated With Histologically Derived Human Ovarian Non-growing Follicle Count in Reproductive Age Women

STUDY QUESTION: Are lifestyle factors (smoking BMI alcohol use and oral contraceptive pill use) associated with the human ovarian reserve as determined by the total ovarian non-growing follicle number? SUMMARY ANSWER: Light to moderate alcohol use was significantly associated with greater ovarian non-growing follicle (NGF) count whereas other lifestyle factors were not significantly related. WHAT IS KNOWN ALREADY: A single previous investigation has suggested that smoking and alcohol use are associated with lower ovarian follicle density. However this investigation utilized follicle density as the outcome of interest rather than the estimated total ovarian NGF count. STUDY DESIGN SIZE DURATION: This cross-sectional investigation included a convenience sample of premenopausal women from two different academic sites the University of Washington (n = 37 from 1999-2004) and the University of Oklahoma (n = 73 from 2004-2013) undergoing incidental oophorectomy at the time of hysterectomy (total n = 110 age range 21-52 years). PARTICIPANTS/MATERIALS SETTING METHODS: Prior to undergoing oophorectomy participants completed detailed questionnaires regarding lifestyle exposures. Following surgery total ovarian NGF counts were determined with systematic random sampling rules and a validated fractionator/optical dissector technique. Associations between lifestyle factors and log-transformed ovarian follicle counts were determined using multivariable linear regression. MAIN RESULTS AND THE ROLE OF CHANCE: After controlling for age BMI oral contraceptive pill (OCP) use tobacco use and site of collection cumulative alcohol use (measured in alcoholic drinks per day multiplied by years of drinking) was associated with ovarian NGF count. Women reporting light (>0 to 3 drink-years) BMI OCP use nor tobacco use were significantly associated with the ovarian NGF count. Similar patterns of association with moderate cumulative alcohol use were observed when evaluating associations with pre-antral follicles and total follicle counts. LIMITATIONS REASONS FOR CAUTION: All participants in this convenience sample had a benign indication for hysterectomy and therefore may not be broadly representative of the population without such an indication. Additionally lifestyle factors were self-reported and the sample size of the present investigation limits our ability to detect associations of smaller magnitude. WIDER IMPLICATIONS OF THE FINDINGS: While our findings are in disagreement with a single investigation that utilized human follicle density as the outcome of interest they are consistent with many studies investigating the relationship between lifestyle factors and the age of spontaneous menopause. Furthermore they suggest a mechanism that does not involve accelerated follicular atresia to explain the association between smoking and an earlier age of menopause. STUDY FUNDING/COMPETING INTERESTS: This investigation was funded by NIA R29-HD37360-04 (N.A.K.) and OCAST HR04-115 (K.R.H.) and by the National Institute of General Medical Sciences Grant 1 U54GM104938 (J.D.P.). There is no conflict of interest. (c) The Author 2015. Published by Oxford University Press on behalf of the European Society of Human Reproduction and Embryology. All rights reserved. For Permissions please email: journals.permissions@oup.com.

Lifestyle factors associated with histologically derived human ovarian non-growing follicle count in reproductive age women.

Are lifestyle factors (smoking, BMI, alcohol use and oral contraceptive pill use) associated with the human ovarian reserve as determined by the total ovarian non-growing follicle number? Light to moderate alcohol use was significantly associated with greater ovarian non-growing follicle (NGF) count, whereas other lifestyle factors were not significantly related. A single previous investigation has suggested that smoking and alcohol use are associated with lower ovarian follicle density. However, this investigation utilized follicle density as the outcome of interest rather than the estimated total ovarian NGF count. This cross-sectional investigation included a convenience sample of premenopausal women from two different academic sites, the University of Washington (n = 37, from 1999-2004) and the University of Oklahoma (n = 73, from 2004-2013), undergoing incidental oophorectomy at the time of hysterectomy (total n = 110, age range 21-52 years). Prior to undergoing oophorectomy, participants completed detailed questionnaires regarding lifestyle exposures. Following surgery, total ovarian NGF counts were determined with systematic random sampling rules and a validated fractionator/optical dissector technique. Associations between lifestyle factors and log-transformed ovarian follicle counts were determined using multivariable linear regression. After controlling for age, BMI, oral contraceptive pill (OCP) use, tobacco use and site of collection, cumulative alcohol use (measured in alcoholic drinks per day multiplied by years of drinking) was associated with ovarian NGF count. Women reporting light (>0 to <1 drink-years) and moderate (1-3 drink-years) alcohol use had greater NGF counts (β = 0.75, P = 0.04, and β = 1.00, P = 0.03; light and moderate use, respectively) as compared with non-users. Neither heavier alcohol use (>3 drink-years), BMI, OCP use, nor tobacco use were significantly associated with the ovarian NGF count. Similar patterns of association with moderate cumulative alcohol use were observed when evaluating associations with pre-antral follicles and total follicle counts. All participants in this convenience sample had a benign indication for hysterectomy, and therefore may not be broadly representative of the population without such an indication. Additionally, lifestyle factors were self-reported, and the sample size of the present investigation limits our ability to detect associations of smaller magnitude. While our findings are in disagreement with a single investigation that utilized human follicle density as the outcome of interest, they are consistent with many studies investigating the relationship between lifestyle factors and the age of spontaneous menopause. Furthermore, they suggest a mechanism that does not involve accelerated follicular atresia to explain the association between smoking and an earlier age of menopause. This investigation was funded by NIA R29-HD37360-04 (N.A.K.) and OCAST HR04-115 (K.R.H.) and by the National Institute of General Medical Sciences, Grant 1 U54GM104938 (J.D.P.). There is no conflict of interest.

Pyramidal Cell Loss in the Cornu Ammonis of 32-day-old Female Rats Following Exposure to a 900 Megahertz Electromagnetic Field During Prenatal Days 13–21

The number of studies reporting that the electromagnetic field (EMF) emitted by mobile phones affects human health is increasing by the day. In previous studies we reported that a 900 megahertz (MHz) EMF applied throughout the prenatal period reduced the number of pyramidal cells in the cornu ammonis of rat pups in the postnatal period. In this study we investigated the effect of a 900 MHz EMF applied on days 13-21 of the prenatal period on the number of pyramidal cells in the cornu ammonis of rat pups in the postnatal period. For that purpose, pregnant rats were divided into experimental and control groups. Experimental group pregnant rats were exposed to the effect of a 900 MHz EMF on days 13-21 of pregnancy. No procedure was applied to the control group. Newborn female rat pups were added to the study, and no procedure was performed on these after birth. Five newborn female rats were obtained from the experimental group and six from the control group. All female rat pups were decapitated on the postnatal 32nd day, and histological procedures were performed on the brain tissues. Sections were stained with Cresyl fast violet. The optical dissector technique was used to estimate the total number of pyramidal cells in the cornu ammonis. Sections of cornu ammonis were subjected to histopathological evaluations. Our results showed that exposure to 900 MHz EMF during prenatal days 13-21 led to a significant decrease in the number of pyramidal cells in the cornu ammonis of the experimental group female rat pups (P<0.05). Histopathological examination revealed picnotic cells in the cornu ammonis in experimental female rat pups. The pyramidal cell loss in the cornu ammonis may therefore be attributed to exposure to 900 MHz EMF in days 13-21 of the prenatal period. NeuroQuantology | December 2013 | Volume 11 | Issue 4 | Page 591-599

Neuronal loss in the caudal intralaminar thalamic nuclei in a primate model of Parkinson’s disease

In light of postmortem human studies showing extensive degeneration of the center median (CM) and parafascicular (Pf) thalamic nuclei in Parkinson's disease patients, the present study assessed the extent of neuronal loss in CM/Pf of non-human primates that were rendered parkinsonian by repeated injections of low doses of 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP). In order to determine the course of CM/Pf degeneration during the MPTP intoxication, motor-asymptomatic animals with partial striatal dopamine denervation were also used. The Cavalieri's principle for volume estimation and the unbiased stereological cell count method with the optical dissector technique were used to estimate the total number of neurons in the CM/Pf. We found substantial neurons loss in the CM/Pf in both, motor-symptomatic MPTP-treated monkeys in which the striatal dopamine innervation was reduced by more than 80%, and in motor-asymptomatic MPTP-treated animals with 40-50% striatal dopamine loss. In MPTP-treated parkinsonian monkeys, 60 and 62% neurons loss was found in CM and Pf, respectively, while partially dopamine-depleted asymptomatic animals displayed 59 and 52% neurons loss in the CM and Pf, respectively. Thus, our study demonstrates that the CM/Pf neurons loss is an early phenomenon that occurs prior to the development of parkinsonian motor symptoms in these animals. In contrast, the neighboring mediodorsal nucleus of the thalamus was only mildly affected (18% neurons loss) in the parkinsonian monkeys. Together with recent findings about the possible role of the CM/Pf-striatal system in cognition, our findings suggest that the pathology of the thalamostriatal system may precede the development of motor symptoms in PD, and may account for some of the cognitive deficits in attentional set-shifting often seen in these patients. Future studies in this animal model, and in monkeys with selective lesion of CM or Pf, are needed to further elucidate the role of the CM/Pf-striatal system in normal and parkinsonian conditions.

Effects of electromagnetic radiation on spatial memory and synapses in rat hippocampal CA1.

In this study, we investigated the effects of mobile phone radiation on spatial learning, reference memory, and morphology in related brain regions. After the near-field radiation (0.52–1.08 W/kg) was delivered to 8-week-old Wistar rats 2 hours per day for 1 month, behavioral changes were examined using the Morris water maze. Compared with the sham-irradiated rats, the irradiated rats exhibited impaired performance. Morphological changes were investigated by examining synaptic ultrastructural changes in the hippocampus. Using the physical dissector technique, the number of pyramidal neurons, the synaptic profiles, and the length of postsynaptic densities in the CA1 region were quantified stereologically. The morphological changes included mitochondrial degenerations, fewer synapses, and shorter postsynaptic densities in the radiated rats. These findings indicate that mobile phone radiation can significantly impair spatial learning and reference memory and induce morphological changes in the hippocampal CA1 region.

Neuroprotective effects of Equisetum telmateia in rat

Equisetum telmateia (Equisetaceae) seems to have anti inflammatory and antioxidant properties. In the present study, the neuroprotective effects of organic and inorganic silica were investigated on spinal cord alpha motoneuron of rats after injury of sciatic nerve. After highly compression of sciatic nerve in 42 Wistar rats, the injured rats were divided into sham (n= 6) and two experimental groups which each were divided into 3 subgroups (n= 6). The first subgroups received 3, 6 or 9 injections (15 mg/kg/injection, ip) of horse tail extract and the second subgroups received 3, 6 or 9 injections (6 mg/kg/injection, ip) of sodium meta silicate, respectively. The first injection was made after sciatic nerve injury and the others by 72 hours intervals. After a month, the rats were sacrificed and their spinal cord lumber segment sampled, processed for histological preparation and analyzed stereologicaly (the dissector technique) for estimation of numerical density of alpha motoneurons. The results showed significant decrease in the numerical density of alpha motoneurons in shams (p< 0.05) and no significant differences between experimental and control groups. This may suggest the neuroprotective effects of silica on the survival of alpha motoneurons.

Complementary techniques for unbiased stereology of brain ultrastructure.

Estimating synaptic density is central to any examination of synaptic change. Barring techniques involving exhaustive sectioning, the traditional method for this estimation is the double dissector technique, which derives a ratio of synapses per neuron. This ratio, however, may not always equate to an absolute number of synapses. In contrast, the unbiased brick, while free of these problems, is limited by changes in reference space and cannot provide information on synaptic connectivity. Thus, the ideal analysis is to derive both synaptic measures. When these measures agree, one can discount changes in extraneous factors influencing synaptic counts.

Antral follicle count by transvaginal ultrasound is reflective of the actual primordial follicle pool

Antral follicle count by transvaginal ultrasound is reflective of the actual primordial follicle pool

Patients with vascular dementia due to microvascular pathology have significant hippocampal neuronal loss

Background: Alzheimer's disease (AD) is characterised by functional impairment, cerebral atrophy, and degeneration of specific neuronal populations, especially pyramidal neurones of the cerebral cortex and hippocampal formation. Although patients with...

Primary sensory neurons and satellite cells after peripheral axotomy in the adult rat: timecourse of cell death and elimination.

The timecourse of cell death in adult dorsal root ganglia after peripheral axotomy has not been fully characterised. It is not clear whether neuronal death begins within 1 week of axotomy or continues beyond 2 months after axotomy. Similarly, neither the timecourse of satellite cell death in the adult, nor the effect of nerve repair has been described. L4 and L5 dorsal root ganglia were harvested at 1-14 days, 1-6 months after sciatic nerve division in the adult rat, in accordance with the Animals (Scientific Procedures) Act 1986. In separate groups the nerve was repaired either immediately or following a 1-week delay, and the ganglia were harvested 2 weeks after the initial transection. Microwave permeabilisation and triple staining enabled combined TUNEL staining, morphological examination and neuron counting by the stereological optical dissector technique. TUNEL-positive neurons, exhibiting a range of morphologies, were seen at all timepoints (peak 25 cells/group 2 weeks after axotomy) in axotomised ganglia only. TUNEL-positive satellite cell numbers peaked 2 months after axotomy and were more numerous in axotomised than control ganglia. L4 control ganglia contained 13,983 (SD 568) neurons and L5, 16,285 (SD 1,313). Neuron loss was greater in L5 than L4 axotomised ganglia, began at 1 week (15%, P=0.045) post-axotomy, reached 35% at 2 months ( P<0.001) and was not significantly greater at 4 months or 6 months. Volume of axotomised ganglia fell to 19% of control by 6 months ( P<0.001). In animals that underwent nerve repair, both the number of TUNEL-positive neurons and neuron loss were reduced. Immediate repair was more protective than repair after a 1-week delay. Thus TUNEL positivity precedes actual neuron loss, reflecting the time taken to complete cell death and elimination. Neuronal death begins within 1 day of peripheral axotomy, the majority occurs within the first 2 months, and limited death is still occurring at 6 months. Neuronal death is modulated by peripheral nerve repair and by its timing after axotomy. Secondary satellite cell death also occurs, peaking 2 months after axotomy. These results provide a logical framework for future research into neuronal and satellite cell death within the dorsal root ganglia and provide further insight into the process of axotomy induced neuronal death.

Leukemia inhibitory factor by systemic administration rescues spinal motor neurons in the SOD1 G93A murine model of familial amyotrophic lateral sclerosis

Leukemia inhibitory factor (LIF) is a survival factor for motoneurons. In this study we investigated whether intense systemic LIF therapy prevents the loss of lumbar motoneurons in the transgenic SOD1 G93A mouse model of familial amyotrophic lateral sclerosis. Treatment involved daily 25 μg/kg intraperitoneal injection for a period of 6 weeks starting at 70 days of age. Using the unbiased optical dissector technique, significant rescue of motoneurons in the LIF-treated group (3809±455) was found compared to the vehicle group (1085±140).

Analysis of the testicular phenotype of the follicle-stimulating hormone beta-subunit knockout and the activin type II receptor knockout mice by stereological analysis.

This study evaluated the role of FSH and activin A on testicular function using quantitative stereological analysis of testicular cell types in mice with targeted disruption of genes encoding the FSH beta-subunit and the activin type IIA receptor (ActRIIA). Using the optical dissector technique, the numbers of Sertoli cells and germ cells per testis were determined. Testis weights in homozygous males lacking the FSHbeta gene or the ActRIIA gene were decreased approximately 60% compared with wild-type or respective heterozygotes. Sertoli cell numbers decreased in both homozygous mice by 30-39%, and there was a comparable decline in germ cell numbers in both models. The degree of germ cell attrition increased in the later stages of spermatogenesis from a 46% reduction of spermatogonia to a 60% decrease in round spermatids. As the FSH levels are decreased in both models, the cellular lesion in both is most likely due to the FSH deficiency. Although the decrease in the Sertoli cell complement represents one cause of lower germ cell numbers, the ability of Sertoli cells to nurture germ cells is compromised by the lower FSH levels, as shown by a decrease in the round spermatid to Sertoli cell ratios in both homozygous models. We conclude that the defects in FSH beta-subunit gene knockout and ActRIIA knockout mice are related to diminished FSH action on both Sertoli cell proliferation and the capacity of Sertoli cells to nurture germ cells.

Variation in hippocampal neuron number with age and brain volume.

Hippocampal size and neuron number are reduced in a number of conditions, including temporal lobe epilepsy and Alzheimer's disease. Furthermore, a decrease with advancing age has also been suggested. The present study examined the entire hippocampal formation of 12 subjects aged from 46 to 85 years and free from neurological disease. The volume of seven subregions (CA1, CA2-3, CA4, dentate gyrus, subiculum, presubiculum and white matter) was determined and the number of neurons estimated in each of these grey matter subregions using the optical dissector technique. There was a significant relationship between CA1 neuron number and cerebrum volume. Multivariate analysis showed the greater contribution to the variance in CA1 neuron number was made by cerebrum volume (69%) rather than age (2%) or sex (1%). The findings of this study show that, in neurologically normal individuals, brain size is a major determinant of the number of CA1 neurons.

Hair cell recovery in the chinchilla crista ampullaris after gentamicin treatment: A quantitative approach

The mechanisms for hair cell recovery were investigated after intraotic application of 50 μg gentamicin into the perilymphatic space of the superior semicircular canal of the chinchilla. Histologic evaluation of one normal group and four posttreatment groups (7, 14, 28, and 56 days) was made with light and transmission electron microscopic techniques. The numeric changes of hair cells and supporting cells was quantified with the dissector technique. At 7 and 14 days after treatment, no type I hair cells were present, and 85% and 88% of type II hair cells were lost. Supporting cells decreased to 76% at 7 days, but they recovered to 91% at 14 days. Recovery of the epithelia was evident 28 days after treatment; 83% were type II hair cells, and 3% were type I hair cells. The supporting cell number remained close to normal (86%). Between 14 and 28 days after treatment, there was an increase of 1758 of type II hair cells, representing approximately 125 new hair cells per day. At the same time interval the number of supporting cells remained near normal. These results suggest that new hair cells might be the result of supporting cell mitotic division and differentiation. (Otolaryngol Head Neck Surg 1998;119:255-62.)

Quantification of the process of hair cell loss and recovery in the chinchilla crista ampullaris after gentamicin treatment

The degree of ototoxic drug sensitivity and hair cell repair was determined in the chinchilla horizontal crista ampullaris after intraotic administration of gentamicin. Histological evaluation was made of 22 cristae ampullaris from one normal and six post-treatment (PT) animal groups killed at 1, 4, 7, 14, 28, and 56 days. New hair cell production was quantified, using the dissector technique. Transmission electron microscopy was used to investigate the ultrastructural characteristics of the hair cells in the regenerated epithelium. At I day PT, type I and II hair cells presented cytoplasmic vacuolization, swollen nerve calyces and 20% of type I and 18% of type II hair cells were lost. At 4days PT, 95% of type I hair cells and 14% of type II hair cells had disappeared. In addition, most of the type II hair cells showed clumping of nuclear material. Nerve fibers were not found in the sensory epithelium, but were still observed below the basal lamina. Supporting cells appeared unaffected, maintaining their location in the crista. At 1 and 4 days PT, the damage to hair cells was more pronounced in the central region of the crista ampullaris. The degree of ototoxic damage at 7 days was similar to that of 14 days: no type I hair cells were present and most of the type II hair cells had disappeared; supporting cell nuclei began to occupy the apical part of the sensory epithelium and most of the nerve fibers had retracted. Quantitatively, 87 and 93% of type II hair cells were lost at 7 and 14 days PT, respectively. Initial signs of hair cell recovery began at 28 days PT; immature type II-like hair cells appeared, supporting cell nuclei began to align at the base of the sensory epithelium and nerve fibers penetrating the basal lamina were observed. No type I hair cells were found, but 40% of the normal number of type II hair cells were present. Hair cells appeared to regenerate in the peripheral areas of the cristae ampullaris first. At 56 days PT, an increase in the number of mature type II hair cells was present, supporting cells were aligned at the base of the epithelium, and more nerve fibers appeared to penetrate the basal lamina to the sensory epithelium. Although type I hair cells were absent from the epithelium, 55% of the normal number of type II hair cells were present. At this time, more regenerated hair cells were located in the center of the cristae ampullaris as compared to the periphery. At the transmission electron microscopic level, type II hair cells at different stages of maturation were observed. Some exhibited mature stereocilia, a cuticular plate, and terminal endings with synaptic specialization opposing these hair cells. In conclusion, type I hair cells were more sensitive than type II hair cells to gentamicin intoxication (as they disappeared as early as 4days PT). After 56 days PT, the number of type II hair cells reached 55% of normal. No type I hair cells had regenerated at this time. These results demonstrate quantitatively the differential ototoxic sensitivity and regenerative capacity of hair cells.