Naohito Hato

Bell palsy and herpes simplex virus : identification of viral DNA in endoneurial fluid and muscle

Bell palsy is the most common cause of facial paralysis worldwide; it has an incidence of 20 to 30 per 100 000 persons [1]. Although the second most frequent cause of facial paralysis, the Ramsay-Hunt syndrome, is known to be caused by reactivated varicella-zoster virus [2], the etiologic agent responsible for Bell palsy has not been identified. Many events, such as viral infection [1, 3, 4], ischemia [5], and autoimmune reaction [6], have been proposed as causes of Bell palsy. Viral infection is thought to be the most likely cause [7]. However, it is rare to find a diagnostic fourfold increase in specific viral antibody titer in the acute and convalescent serum specimens of patients with Bell palsy [3, 4, 7]. Postmortem histopathologic studies of the facial nerve suggest viral neuritis [8], but electron microscopic studies have failed to detect specific viral particles in the facial nerve [9]. Because the etiologic agent of Bell palsy is unknown, treatment of this condition is empiric, varying from observation alone to the use of steroids, surgical decompression, and antiviral agents. We analyzed the viral genomes of herpes simplex virus type 1 (HSV-1), varicella-zoster virus, and Epstein-Barr virus using polymerase chain reaction (PCR) on facial nerve endoneurial fluid specimens and specimens of posterior auricular muscle innervated by the facial nerve. Methods Patients and Specimens During a 4-year period, 14 of 170 patients with Bell palsy and 9 of 51 patients with the Ramsay-Hunt syndrome had decompression surgery 12 to 87 days after the onset of facial palsy. None had benefited from medical management. All patients and controls gave informed consent. Two types of clinical specimens were collected intraoperatively: endoneurial fluid from the facial nerve and tissue from the posterior auricular muscle. A piece of auricular muscle was resected after skin incision, and we obtained endoneurial fluid by absorbing it with a small, sterilized surgical sponge held at the mastoid segment immediately after the epineural sheath was incised. We stored both specimens immediately at 80 C and continued to store them at that temperature until PCR analysis was done. Control specimens of endoneurial fluid and posterior auricular muscle were collected during decompression surgery from four patients with temporal bone fracture or bacterial infection concomitant with otitis media. Posterior auricular muscle specimens were obtained during tympanoplasty from five patients with chronic otitis media who did not have facial paralysis. As an additional control, a piece of neural tissue was obtained from each of three patients with parotid tumors or facial neuroma whose facial nerves had already been affected (Table 1). Table 1. Clinical Data and Polymerase Chain Reaction Results in Patients with Bell Palsy, Patients with the Ramsay-Hunt Syndrome, and Controls* Polymerase Chain Reaction To amplify and identify the HSV, varicella-zoster virus, and Epstein-Barr virus genomes, five sets of virus-specific primers and internal oligonucleotide probes were synthesized for PCR and Southern blot analysis. Primer set 1 was prepared for amplification of the HSV-1 genome, which is located on the US6 gene [10]. Primer set 2 was designed to amplify both HSV-1 and HSV-2. A sense primer (5-CCACCGAGCGGCAGGTGATC-3) and an antisense primer (5-GCCGACCGCCTGCTCGTGCT-3) are located on the UL44-45 gene [11]. Using primer set 2, we discriminated between HSV-1 (578 base pairs) and HSV-2 (621 base pairs) on the basis of size. Nucleotide sequences of the HSV specific internal probe are 5-GAGGCGATCGAGTGGGT-3. Primer sets 3 and 4 were prepared for varicella-zoster virus amplification (genes 29 and 62, respectively) [12], and primer set 5 was prepared for Epstein-Barr virus amplification (latent cycle gene) [13]. The sensitivities of the primer sets were assessed by making serial 10-fold dilutions of each purified DNA sample. The limits of detection for primer sets 1, 2, 3, 4, and 5 were about 10, 100, 10, 100, and 10 femtograms, respectively. Samples of endoneurial fluid (about 10 L), posterior auricular muscle (2 mg to 5 mg), and nerve tissue (0.5 mg to 1 mg) were completely digested with proteinase K. Polymerase chain reaction amplification and subsequent hybridization with Southern blot analysis were done as described previously [14]. Rigid precautions against contamination in the sample processing included the use of water controls replacing the DNA samples in all amplifications. Serum Antibody Titers We examined serum antibody titers by using the complement fixation test for HSV-1 and varicella-zoster virus and by using fluorescent antibody methods for Epstein-Barr virus 8 to 38 days after the onset of facial paralysis. Results We amplified HSV-1-specific DNA fragments from the US6 and UL44-45 genes in both endoneurial fluid and posterior auricular muscle specimens obtained from patients with Bell palsy. The PCR-amplified products of the US6 gene were detected by Southern blot analysis in 10 of the 13 fluid specimens (77%) and 8 of the 14 muscle specimens (57%); the products of the UL44-45 gene were detected in 4 of the 13 fluid specimens (31%) and 7 of the 14 muscle specimens (50%). Neither varicella-zoster virus nor Epstein-Barr virus was detected in the same clinical specimens (Figure 1, top; (Table 1). We did not detect HSV-1 DNA in either the fluid or the muscle specimens of three patients with Bell palsy [patients 9, 12, and 13]. The PCR-amplified DNA fragments of the US6 gene from two patients with Bell palsy (patients 6 and 7) were sequenced directly after asymmetric PCR was done as described previously [14]. The nucleotide sequences of the amplified products (221 base pairs) were identical to those of the HSV-1 genome submitted to the GenBank (Mountain View, California [data bank with genetic information]) with accession numbers J02217 and K02372. Figure 1. Amplification of herpes simplex virus type 1 (HSV-1) and varicella-zoster virus (VZV) genomes from clinical samples. top bottom middle We detected varicella-zoster virus DNA from gene 29 or gene 62 in specimens obtained from patients with the Ramsay-Hunt syndrome only (Figure 1, middle; Table 1). Gene 29 was detected in 8 of the 9 patients (89%), and gene 62 was detected in 6 of the 9 patients (67%). On the other hand, we could not amplify HSV-1, varicella-zoster virus, or Epstein-Barr virus DNA from any specimens obtained from the other controls (Figure 1, bottom; Table 1). Serum antibody titer to HSV-1 was positive in 12 of 13 patients (92%) with Bell palsy, in 4 of 9 patients (44%) with the Ramsay-Hunt syndrome, and in 5 of 9 controls (56%) (Table 1). The prevalence of HSV-1 antibody in patients with Bell palsy was significantly higher than that in controls (P < 0.05, Fisher exact test). However, antibody titers to HSV-1 in patients with Bell palsy were not significantly higher than those of controls, as previously reported [3, 4]. Discussion We found HSV-1 DNA in 11 of 14 patients (79%) with Bell palsy, and we found varicella-zoster virus DNA in 8 of the 9 patients (89%) with the Ramsay-Hunt syndrome. The identification of viral DNA may not always be definitive evidence that a particular agent causes a disease process, because PCR can amplify viral DNA regardless of whether the virus is in the infective, lytic, or latent state. The presence of latent HSV-1 and varicella-zoster virus genomes has also been shown by PCR in the geniculate ganglion of human facial nerves at autopsy [15-17]. However, HSV-1 and varicella-zoster virus usually remain dormant in ganglia and would probably not be detected in the endoneurial fluid or auricular muscle unless they were reactivated. This hypothesis was supported by our inability to detect either HSV-1 or varicella-zoster DNA in controls. Cranial nerve surgery often reactivates latent HSV, causing labial and facial herpetic lesions 48 to 72 hours after surgery [18]. However, because we obtained specimens within 2 hours of beginning decompression surgery, reactivation of the virus in the muscle or fluid was probably not induced by surgery. If this surgery did reactivate latent HSV-1, viral DNA should also have been detected in patients with the Ramsay-Hunt syndrome and in other controls who were seropositive for HSV-1. Triggers known to be associated with Bell palsy are also known to reactivate HSV. Preceding stress, such as upper respiratory tract infection, fever, dental extraction, menstruation, or exposure to cold might reactivate latent HSV-1 in the geniculate ganglion. After the virus reactivates, it destroys ganglion cells and spreads into the endoneurial fluid. The virus also infects Schwann cells, leading to demyelinization and inflammation of the facial nerve [19]. This inflammatory response has been shown by gadolinium-enhanced magnetic resonance imaging in patients with Bell palsy and in patients with the Ramsay-Hunt syndrome [20]. Given the known neuropathogenicity of HSV-1 and the presence of HSV-1 DNA in the lesional site of the facial nerve specific to patients with Bell palsy, we conclude that HSV-1 infection in the facial nerve is directly related to the pathogenesis of Bell palsy just as the varicella-zoster virus is directly related to the pathogenesis of the Ramsay-Hunt syndrome. There are two possible explanations for our failure to detect HSV-1 in three of the patients with Bell palsy [patients 9, 12, and 13]: 1) the limited sensitivity of PCR analysis to detect small amounts of viral DNA and 2) the presence of an etiologic agent other than HSV-1. More data are required to determine the percentage of patients with Bell palsy in whom HSV-1 is the etiologic agent of Bell palsy, but our findings suggest that HSV-1 infection is the major cause of Bell palsy and that treatment with appropriate antiviral agents might benefit most patients with this condition. Drs. Mizobuchi, Nakashiro, and Doi: Department of Neuropsychiatry, Ehime Universit

Induction of myasthenia by immunization against muscle-specific kinase

Muscle-specific kinase (MuSK) is critical for the synaptic clustering of nicotinic acetylcholine receptors (AChRs) and plays multiple roles in the organization and maintenance of neuromuscular junctions (NMJs). MuSK is activated by agrin, which is released from motoneurons, and induces AChR clustering at the postsynaptic membrane. Although autoantibodies against the ectodomain of MuSK have been found in a proportion of patients with generalized myasthenia gravis (MG), it is unclear whether MuSK autoantibodies are the causative agent of generalized MG. In the present study, rabbits immunized with MuSK ectodomain protein manifested MG-like muscle weakness with a reduction of AChR clustering at the NMJs. The autoantibodies activated MuSK and blocked AChR clustering induced by agrin or by mediators that do not activate MuSK. Thus MuSK autoantibodies rigorously inhibit AChR clustering mediated by multiple pathways, an outcome that broadens our general comprehension of the pathogenesis of MG.

Efficacy of early treatment of Bell's palsy with oral acyclovir and prednisolone

Objective To investigate the therapeutic effects of acyclovir and prednisolone in relation to the timing of treatment in Bells palsy. Study Design This was a retrospective study of 480 Bells palsy patients who were treated with oral acyclovir and prednisolone (94 cases) or prednisolone alone (386 cases). Patients Patients met the after criteria: (1) severe or complete Bells palsy with a score lower than 20 on the 40-point Yanagihara facial score and (2) treatment started within 7 days after onset. The patients were treated with oral prednisolone (60–40 mg/day) with or without oral acyclovir (2,000 mg/day). Main Outcome Measure Rate of recovery, which was defined as a facial score of 36 or more, and the absence of contracture with synkinesis. Results The overall recovery rate of patients treated with acyclovir and prednisolone was 95.7 percent, which was better than that of patients treated with prednisolone alone (88.6%). The recovery rate in patients who began the combined therapy within 3 days of the onset of palsy was 100 percent and early treatment resulted in early remission. In contrast, the recovery rate in patients who started the combined therapy more than 4 days after onset was 86.2 percent. Conclusion These results suggest that early diagnosis and treatment within 3 days of the onset of paralysis are necessary for maximal efficacy of combined acyclovir and prednisolone therapy for Bells palsy.

Ramsay Hunt syndrome in children

In a retrospective study, 52 children were diagnosed with Ramsay Hunt syndrome. The facial palsy was milder and complete recovery of the function was achieved in 78.6% of patients. Associated cranial neuropathies were less common in children than in adults. The timing of vesicle appearance tended to be delayed in children. In preschool children, Ramsay Hunt syndrome was rare, although the frequency has recently increased. The syndrome is relatively common in older children. This study suggested that vaccination can prevent or reduce the occurrence of Ramsay Hunt syndrome. Ann Neurol 2000;48:254–256

Transmastoid decompression as a treatment of Bell palsy

OBJECTIVE: We sought to assess the efficacy of transmastoid decompression after steroid treatment. STUDY DESIGN: One hundred one adults with Bell palsy having denervation exceeding 95% after steroid treatment were divided into 2 groups. In 58 patients decompression from the labyrinthine segment to the stylomastoid foramen was performed, and the remaining 43 patients were only followed up. Using the Yanagihara score and House Brackmann grading system, the recovery from the palsy was assessed. RESULTS: There was a statistically significant difference in the final facial score of the 2 groups. Within 60 days after the onset, the chance of better recovery from the palsy was higher in the patients with decompression. CONCLUSION: In the era of steroid treatment, we cannot discard the transmastoid decompression of the facial nerve in the treatment of severe Bell palsy with profound denervation, although further effort is needed to obtain definitive evidence to show the benefit of the operation.

Varicella-Zoster Virus Distribution in Ramsay Hunt Syndrome Revealed by Polymerase Chain Reaction

The pathogenesis of facial nerve paralysis and vestibulo-cochlear dysfunction of Ramsay Hunt syndrome remains unclear as varicella-zoster virus (VZV) has not been demonstrated in the lesions. Using the polymerase chain reaction, we detected VZV genomes not only in the vesicles on the auricles or oral cavity but also in the facial nerve sheath, middle ear mucosa and cerebrospinal fluid from patients with Ramsay Hunt syndrome. The VZV genome was undetectable in the same kinds of clinical samples obtained from control patients with facial nerve paralysis of other etiologies. The results indicated that VZV spreads widely in the neural components, mucocutaneous tissue and cerebrospinal fluid. The present study will facilitate better understanding of the pathogenesis of facial nerve paralysis, vertigo, hearing impairment and other cranial nerve dysfunction of Ramsay Hunt syndrome.

A randomized controlled clinical trial of topical insulin-like growth factor-1 therapy for sudden deafness refractory to systemic corticosteroid treatment

BackgroundTo date, no therapeutic option has been established for sudden deafness refractory to systemic corticosteroids. This study aimed to examine the efficacy and safety of topical insulin-like growth factor-1 (IGF-1) therapy in comparison to intratympanic corticosteroid therapy.MethodsWe randomly assigned patients with sudden deafness refractory to systemic corticosteroids to receive either gelatin hydrogels impregnated with IGF-1 in the middle ear (62 patients) or four intratympanic injections with dexamethasone (Dex; 58 patients). The primary outcome was the proportion of patients showing hearing improvement (10 decibels or greater in pure-tone average hearing thresholds) 8 weeks after treatment. The secondary outcomes included the change in pure-tone average hearing thresholds over time and the incidence of adverse events.ResultsIn the IGF-1 group, 66.7% (95% confidence interval [CI], 52.9-78.6%) of the patients showed hearing improvement compared to 53.6% (95% CI, 39.7-67.0%) of the patients in the Dex group (P = 0.109). The difference in changes in pure-tone average hearing thresholds over time between the two treatments was statistically significant (P = 0.003). No serious adverse events were observed in either treatment group. Tympanic membrane perforation did not persist in any patient in the IGF-1 group, but did persist in 15.5% (95% CI, 7.3-27.4%) of the patients in the Dex group (P = 0.001).ConclusionsThe positive effect of topical IGF-1 application on hearing levels and its favorable safety profile suggest utility for topical IGF-1 therapy in patients with sudden deafness.Trial registrationUMIN Clinical Trials Registry Number UMIN000004366, October 30th, 2010.

Protection against ischemic cochlear damage by intratympanic administration of AM-111.

Objective AM-111, a cell-permeable peptide inhibitor of c-Jun N-terminal kinase, was investigated for its protective effects against ischemic damage of the cochlea in gerbils. Methods Transient cochlear ischemia was introduced in animals by occluding the bilateral vertebral arteries for l5 minutes. Then, 10 &mgr;l of AM-111 at a concentration of l, 10, or 100 &mgr;M in hyaluronic acid gel formulation was applied onto the round window 30 minutes after the insult. Gel without active substance was used in a control group. Treatment effects were evaluated by auditory brainstem response (ABR) and histology of the inner ear. Results In controls, transient cochlear ischemia caused a 25.0 ± 5.0 dB increase in the ABR threshold at 8 kHz and a decrease of 13.3 ± 2.3% in inner hair cells at the basal turn on Day 7. Ischemic damage was mild at 2 and 4 kHz. When the animals were treated with AM-111 at 100 &mgr;M, cochlear damage was significantly reduced: the increase in ABR threshold was 3.3 ± 2.4 dB at 8 kHz, and the inner hair cell loss was 3.1 ± 0.6% at the basal turn on Day 7. The effects of AM-111 were concentration dependent: 100 &mgr;M was more effective than 1 or 10 &mgr;M. Conclusion Direct application of AM-111 in gel formulation on the round window was effective in preventing acute hearing loss because of transient cochlear ischemia.

Edematous Swelling of the Facial Nerve in Bell's Palsy

Surgical decompression of the intratemporal facial nerve from the geniculate ganglion to the stylomastoid foramen was carried out in 91 patients with Bells palsy. All of the patients had denervation exceeding 95%, and a suprastapedial lesion. Edematous swelling of the nerve was assessed using the following three grades: + +, nerve swells beyond the bony facial canal; +, nerve swells beyond the nerve sheath, but within the bony canal, and -, no notable swelling observed. Varying degrees of swelling of the nerve were noted in all of the patients from onset to the end of the ninth week. The incidence of + + swelling was highest within 3 weeks of onset and then declined. + + swelling was most often noted in the vicinity of the geniculate ganglion, and was thought to be a manifestation of inflammation due to herpes simplex virus infection. There was a clear time dependency of the swelling in the horizontal and pyramidal segments, but not in the mastoid segment. After the ninth week, the incidence of swelling decreased sharply and no swelling of the nerve was observed in about one-third of the patients. Considering the etiology and the analysis of edematous swelling, we propose that the course of Bells palsy be differentiated into an acute phase (the first 3 weeks after onset), a subacute phase (from the fourth to ninth weeks) and a chronic phase (after the tenth week).Surgical decompression of the intratemporal facial nerve from the geniculate ganglion to the stylomastoid foramen was carried out in 91 patients with Bell’s palsy. All of the patients had denervation exceeding 95% and a suprastapedial lesion. Edematous swelling of the nerve was assessed using the following three grades: + + , nerve swells beyond the bony facial canal; + , nerve swells beyond the nerve sheath, but within the bony canal, and − , no notable swelling observed. Varying degrees of swelling of the nerve were noted in all of the patients from onset to the end of the ninth week. The incidence of + + swelling was highest within 3 weeks of onset and then declined. + + swelling was most often noted in the vicinity of the geniculate ganglion, and was thought to be a manifestation of inflammation due to herpes simplex virus infection. There was a clear time dependency of the swelling in the horizontal and pyramidal segments, but not in the mastoid segment. After the ninth week, the incidence of swelling decreased sharply and no swelling of the nerve was observed in about one-third of the patients. Considering the etiology and the analysis of edematous swelling, we propose that the course of Bell’s palsy be differentiated into an acute phase (the first 3 weeks after onset), a subacute phase (from the fourth to ninth weeks) and a chronic phase (after the tenth week).

Idiopathic sudden sensorineural hearing loss in Japan

Abstract Conclusion: An epidemiological survey of hospitals and private clinics in Japan regarding idiopathic sudden sensorineural hearing loss (SSNHL) revealed that the incidence of SSNHL was 60.9 per 100 000 population. There were more females than males in the younger generation. Objective: The incidence of SSNHL varies largely by country. Because the Japanese criteria for diagnosing SSNHL have changed in accordance with those widely used in other parts of the world, a clinicoepidemiological study was undertaken using the new criteria. Methods: Ehime, Aichi, and Iwate Prefectures were selected from the western, central, and northeastern regions of Japan, respectively. The subjects for this study were patients who suffered SSNHL between April 1, 2012 and March 31, 2013. Questionnaires were mailed to all hospitals and private clinics in which ENT doctors were working. Initial and final audiograms were requested for 10% of the patients. Results: In all, 78 of 90 hospitals (87%) and 303 of 407 private clinics (74%) responded. It was reported that 1663 patients visited hospitals and 3090 patients visited only private clinics. It was estimated that 6205 SSNHL patients visited hospitals or private clinics in 1 year from a population of 10 145 000. Also, 23% of patients suffered acute low-tone SNHL (female to male ratio; 3:1 in definite cases).