A Novel Tumor Staging System Incorporating Tumor Regression Grade (TRG) With Lymph Node Status (ypN-Category) Results in Better Prognostication Than ypTNM Stage Groups After Neoadjuvant Therapy for Esophageal Squamous Cell Carcinoma.

OBJECTIVE: This study aims at constructing a staging system incorporating tumor regression grade and ypN-category (TRG-N) in patients with neoadjuvant therapy before esophagectomy. It is hypothesized that this would prognosticate better than the current American Joint Committee on Cancer (AJCC) postneoadjuvant therapy (ypTNM) stage groups. BACKGROUND: Conventional pathological T-category is defined by the depth of invasion, and may lose prognostic relevance after neoadjuvant therapy. TRG defines treatment response by the degree of tumor regression, and when combined with ypN-category may be more prognostic than AJCC postneoadjuvant therapy (ypTNM) stage groups. METHODS: A training cohort of 210 patients with esophageal squamous cell carcinoma and who had had neoadjuvant therapy before esophagectomy were studied. A validation cohort comprised 107 patients from another hospital. Resected esophagi were assessed by ypT-category and TRG, the latter assigned according to the Becker 4-tier system. These categories were grouped with ypN-category into a TRG-N system. Patients' survival was compared between the current AJCC postneoadjuvant therapy (ypTNM) stage groups and this TRG-N system. RESULTS: In the training cohort, 5-year survival rates according to ypTNM stage I, II, IIIA, IIIB, and IVA were 53%, 39.4%, 47%, 18.3%, and 0%, respectively. For TRG-N stages I, II, III, and IV, the respective figures were 59.6%, 43.5%, 23.8%, and 15.6%. TRG-N stage showed better fit in survival than ypTNM stage groups, indicated by lower Akaike Information Criteria (AIC) and Bayesian Information Criterion values. Similar results were found in the validation cohort. Multivariate analysis showed that TRG-N stage ( P =0.02), age ( P =0.006), and sex ( P =0.005) were independent prognostic factors. CONCLUSION: TRG-N stage shows better prognostication than the AJCC postneoadjuvant therapy (ypTNM) stage groups.

A standardized approach for the thoracic dissection in robotic-assisted minimally invasive esophagectomy (RAMIE).

Robot-assisted minimally invasive esophagectomy (RAMIE) is increasingly being adopted as the preferred surgical treatment for esophageal cancer, as it is superior to open esophagectomy and a good alternative to conventional minimally invasive esophagectomy. This paper addresses the technical details of the thoracoscopic phase of RAMIE, including the operating room set-up, patient positioning, port placement, and surgical steps.

Do dysplastic proximal resection margins predict the risk of anastomotic recurrence and overall survival in patients with esophageal squamous cell carcinoma?

Positive proximal resection margins are strongly associated with anastomotic recurrence in esophageal cancer. However, the prognostic significance of dysplastic proximal resection margins remains unclear. The aim of this study is to investigate whether the dysplastic proximal resection margin can predict anastomotic recurrence and overall survival in patients with esophageal squamous cell carcinoma. Between 2000 and 2014, patients with esophageal squamous cell carcinoma who received a nonpalliative resection and survived the perioperative period were included. Two expert pathologists independently reviewed the proximal resection margin status, which was classified as negative, dysplastic, or positive. The kappa statistic was used to test interobserver reliability. Anastomotic recurrence and overall survival served as the main outcome measures. The study cohort consisted of 469 patients (445 males and 27 females). There was an excellent interobserver agreement for negative (kappa = 0.88), dysplastic (kappa = 0.88), and positive (kappa = 1) proximal resection margins-which were identified in 418 (89.1%), 37 (7.9%), and 14 (3.0%) patients, respectively. After a median follow-up of 21.6 months, 30 (6.4%) patients developed an anastomotic recurrence. Compared with patients with negative proximal resection margins (24/418, 5.7%), the occurrence of anastomotic recurrence was more commonly observed in those with positive proximal resection margins (3/14, 21.4%, P = 0.017) but not in those with dysplastic proximal resection margins (3/37, 8.1%, P = 0.56). Multivariable Cox regression analysis identified positive proximal resection margins (hazard ratio: 5.93, P = 0.010) and advanced clinical stage (hazard ratio: 12.04, P = 0.023) as independent risk factors for anastomotic recurrence. Dysplastic proximal resection margins were not retained in the model as an independent predictor (hazard ratio: 1.38, P = 0.602). The 5-year overall survival rates of patients with negative (38.2%) and dysplastic margins (27.0%) were similar (P = 0.814), and significantly higher than that observed in those with positive proximal resection margins (9.5%, P = 0.015). In conclusion, dysplastic proximal resection margins can be identified in at least 7.9% of patients with esophageal squamous cell carcinoma, but neither they are associated with an increased risk of anastomotic recurrence nor they portend a poor overall survival.

Induction therapy before surgery improves survival in patients with clinical T3N0 esophageal cancer: a nationwide study in Taiwan.

The utility of induction therapy (IT) in patients with resectable esophageal cancer remains controversial, especially when clinical evidence of nodal metastases is lacking. We sought to compare the survival impact of IT versus upfront surgery (US) in patients with cT3N0 esophageal cancer. We searched the Taiwan Cancer Registry for patients with cT3N0 esophageal cancer who underwent US or IT between 2008 and 2013. Multivariate Cox regression analysis was used to analyze the potential benefits of IT in terms of overall survival (OS) and disease-free survival (DFS). Of the 11752 patients with esophageal cancer included in the nationwide database, 762 (6.5%) had cT3N0 disease. Most cases (720 [94.5%]) had a histological diagnosis of squamous cell carcinoma. Of them, 135 received IT (the IT group) and 237 received surgery first (the US group). In the US group, pretreatment clinical staging was accurate in 47.9% of patients. Twenty-one (8.97%) were clinically overstaged (pT1-2N0), whereas 101 (43.17%) were clinically understaged (pT4N0 or pTanyN1-3). The presence of unexpected nodal metastases was identified in 92.1% of clinically understaged patients. In the IT group, 28 (20.74%) patients did not proceed to surgery after IT. The use of IT was associated with higher R0 resection rates and fewer pathological nodal metastases, despite unexpected M1 disease being more common (all P< 0.05). The 5-year OS rate was significantly higher (42%) in the IT group than in the US group (33%, P= 0.032). Similar findings were observed in terms of 5-year DFS (37% in the IT group versus 29% in the US group, P= 0.009). Multivariate analysis identified US (hazard ratio: 1.42, P= 0.03) and non-R0 resection (hazard ratio: 1.58, P= 0.03) as independent adverse prognostic factors. We found that 43.17% of patients with cT3N0 disease undergoing primary surgery had their disease understaged. The use of IT before esophagectomy significantly improves OS and DFS in patients with clinical T3N0 esophageal squamous cell carcinoma.

Perineural invasion through the sheath in posttherapy esophagectomy specimens predicts poor survival in patients with esophageal squamous cell carcinoma.

BACKGROUND: The prognostic impact of perineural invasion (PNI) in patients with esophageal cancer who receive neoadjuvant chemoradiotherapy (nCRT) remains unclear. METHODS: A thorough pathological review of PNI was performed on post-nCRT esophagectomy specimens obtained from non-ypT0 patients with esophageal squamous cell carcinoma (ESCC). When PNI was identified, it was classified according to the presence or absence of penetration through the nerve sheath (i.e., PNI surrounding the nerve sheath [PNI-SS] versus PNI penetrating through the nerve sheath [PNI-TS]). The impact of PNI on overall survival (OS) was assessed in combination with clinical and pathological risk factors. RESULTS: A total of 177 eligible patients were identified between 1998 and 2008. PNI was identified in 43.5% (77/177) of participants. Of them, 33 and 44 had PNI-SS and PNI-TS, respectively. The 5-year OS rate of patients with PNI-TS was significantly lower (6.7%) than that observed in those without PNI (30.6%, P < 0.001). However, the 5-year OS observed in the latter group did not differ significantly from that of patients with PNI-SS (26%, P = 0.68). Multivariate analysis identified PNI-TS (hazard ratio [HR] = 1.965, P = 0.02), LVI (HR = 1.514, P = 0.048), and ypN2 stage (HR = 2.39, P = 0.007) as independent adverse prognostic factors for OS. CONCLUSIONS: The presence of PNI-TS after nCRT is associated with poor survival. A thorough assessment of distinct PNI patterns (i.e., PNI-TS versus PNI-SS) should be part of the routine post-nCRT histopathological work-up of ESCC patients.

Predictive value of nodal maximum standardized uptake value of pretreatment [18F]fluorodeoxyglucose positron emission tomography imaging in patients with esophageal cancer.

We retrospectively reviewed 102 patients with esophageal cancer (97.1% squamous cell carcinoma, 96.1% stage III) received FDG-PET staging and were treated by chemoradiotherapy with or without resection to assess whether the pretreatment [18F]fluorodeoxyglucose positron emission tomography (FDG-PET) maximum standardized uptake value (SUVmax) of the primary tumor and metastatic lymph nodes can predict the prognosis of patients with esophageal cancer. Receiver operating characteristic analysis was performed to find the cutoff values for primary tumor SUVmax and nodal SUVmax. The influence of clinical factors including primary tumor SUVmax and nodal SUVmax on local progression-free survival, nodal progression-free survival (NPFS), distant metastases-free survival (DMFS), and overall survival (OS) were evaluated using univariate and multivariate analyses. A total of 40 patients received esophagectomy after neoadjuvant chemoradiotherapy (trimodality), while 62 patients received definitive chemoradiotherapy (dCRT). The median follow-up was 26.4 months. The SUVmax of primary tumor had no significant predictive value on all outcomes, while the SUVmax of metastatic lymph nodes had predictive value on several outcomes. High nodal SUVmax (≥7) predicted for worse outcomes than low nodal SUVmax (<7) in the patients who received dCRT (two-year DMFS, 17% vs. 92%, P < 0.001; NPFS, 14% vs. 81%, P = 0.001; OS, 21% vs. 50%, P = 0.003), but not in those received trimodality. On multivariate analysis of patients receiving dCRT, nodal SUVmax was the strongest independent predictor on DMFS (hazard ratio [HR] 13.93, P < 0.001), NPFS (HR 3.99, P = 0.026), PFS (HR 2.90, P = 0.003), and OS (HR 3.80, P = 0.001). High pretreatment nodal SUVmax predicts worse treatment outcomes for the patients treated with dCRT.

Prognosis of patients with esophageal squamous cell carcinoma who achieve major histopathological response after neoadjuvant chemoradiotherapy.

BACKGROUND: The purpose of this study was to investigate the prognosis and its predictors in patients with esophageal squamous cell carcinoma (ESCC) who achieve major histopathological response (MaHR) after neoadjuvant chemoradiotherapy (nCRT). METHODS: We examined a total of 187 ESCC patients who achieved MaHR following nCRT and survived the perioperative period. MaHR was defined as either absence or <10% vital residual tumor cells (VRTC) in the resected esophagus without nodal involvement. Univariate and multivariate analyses were used to identify factors significantly associated with overall survival (OS). RESULTS: At the time of analysis, 113 patients (60.4%) were dead (5-year OS = 48%; median survival time = 54.8 months). The amount of VRTC (1-10% versus 0% VRTC; hazard ratio [HR] = 1.9, P < 0.001) and the thoroughness of histopathological examination (standard [≤ 4 tumor blocks] versus thorough [> 4 tumor blocks], HR = 1.57; P = 0.013) were independent predictors of OS in multivariate analysis. A stepwise increase in OS was observed in the following groups: patients with 1-10% VRTC identified by the standard protocol, patients with 1-10% VRTC identified by the thorough protocol, patients with 0% VRTC identified by the standard protocol, and patients with 0% VRTC identified by the thorough protocol (5-year OS rates = 20%, 40%, 50%, and 62%, respectively, P < 0.001). CONCLUSIONS: In ESCC patients who achieve MaHR after nCRT, the presence of microscopical residual disease and the thoroughness of histopathological examination are associated with survival.

Characterization of residual tumours at the primary site in patients with a near pathological complete response after neoadjuvant chemoradiotherapy for oesophageal cancer.

BACKGROUND: A 'surgery as needed' strategy has been proposed for patients with oesophageal cancer who truly achieve a pathological complete response (pCR) following neoadjuvant chemoradiotherapy (nCRT). However, the ability to detect residual disease remains problematic. This study investigated the anatomical locations and pathological characteristics of residual cancer in patients with oesophageal squamous cell carcinoma (SCC) who achieved a near pCR following nCRT. METHODS: Patients with oesophageal SCC who achieved a near pCR after nCRT were eligible. Near pCR was defined as residual cancer in the resection specimen representing less than 10 per cent of the apparent original tumour area. RESULTS: Detailed histopathological reassessment of 76 consecutive patients (mean age 54·4 years) with a near pCR was undertaken. Some 32 patients (42 per cent) with a near pCR had no detectable mucosal lesions. Residual tumour was identified most frequently in the submucosal layer (54, 71 per cent), followed by the mucosa (44, 58 per cent), muscle layer (36, 47 per cent) and adventitia (22, 29 per cent) (P < 0·001). Among patients without ypT1a disease, increasing depth of tumour invasion correlated negatively with the likelihood of mucosal involvement. Of patients with ypT3 disease, 16 of 22 had no detectable cancer located in the mucosa, compared with six of 29 with ypT1b disease (P < 0·001). CONCLUSION: Better tools for predicting pCR are required before considering a 'surgery as needed' approach in the management of oesophageal cancer.

Association between the thoroughness of the histopathological examination and survival in patients with esophageal squamous cell carcinoma who achieve pathological complete response after chemoradiotherapy.

The College of American Pathologists guidelines recommend examining at least four representative tumor blocks for determining pathological T stage in patients with primarily resected esophageal cancer. Whether the same pathological requirements are adequate in patients undergoing esophagectomy following neoadjuvant chemoradiotherapy (nCRT) remains unclear. We hypothesized that current examination protocols may underestimate the presence of microscopical residual disease after nCRT, potentially leading to under-staging. We retrospectively reviewed the records of patients with esophageal squamous cancer (ESCC) who were diagnosed as having pathological complete response (pCR) following nCRT. The thoroughness of the pathological examination in pCR patients was examined using (i) the number of blocks examined in suspicious tumor area (≤4 vs. >4), and (ii) the block quotient (calculated as the pretreatment tumor length divided by the number of blocks examined in suspicious tumor area). A total of 91 patients were enrolled. The mean number of blocks used to confirm pCR was 4.8 (range: 2-14). The 5-year overall survival (OS) and disease-free survival (DFS) in the entire cohort were 55% and 65%, respectively. Multivariate analyses identified the block quotient as the only independent predictor of OS and DFS. Receiver operating characteristic curve analysis indicated an optimal cutoff value of 1.4 for the block quotient. Among the patients who achieved pCR, the 5-year DFS differed significantly between subjects with a low (≤1.4) or high (>1.4) block quotient (76% vs. 47%, respectively, P = 0.03). The block quotient (calculated by the pretreatment tumor length divided by the number of blocks) - which reflects the meticulousness of the histopathological examination for confirming pCR - is associated with survival in ESCC patients.

Long-term outcomes following neoadjuvant chemoradiotherapy in patients with clinical T2N0 esophageal squamous cell carcinoma.

The optimal treatment for patients with local esophageal cancer (cT2N0 disease) has not yet been defined. We sought to determine whether neoadjuvant chemoradiotherapy (CRT) can improve prognosis compared with direct esophagectomy in this patient group. Between 1994 and 2005, patients with cT2N0 esophageal squamous cell carcinoma who underwent either neoadjuvant CRT or surgery as first-line treatment were retrospectively reviewed. We collected information on their demographic characteristics, staging modality, clinical and pathological stages, perioperative course, and survival. The study endpoints included tumor recurrence, disease-specific survival (DSS), and overall survival rate. Of the 71 eligible patients, 14 received an esophagectomy first, whereas the remaining 57 received neoadjuvant CRT first. Despite the high pathological complete response (pCR) rate of 37% after neoadjuvant CRT, routine neoadjuvant CRT did not translate into better survival compared to direct surgery (5-year DSS: 39% vs. 68%, P= 0.17). The dramatic survival difference between pCR and non-pCR patients (5-year DSS: 85% vs. 4%, P < 0.001) accounts for these unsatisfactory results. In our series, the administration of neoadjuvant CRT to patients with clinical stage T2N0 esophageal squamous cell carcinoma did not significantly improve outcomes compared with direct esophagectomy.

Granulation tissue formation following Dumon airway stenting: the influence of stent diameter.

OBJECTIVES: The present study aimed to determine whether stent diameter influences granulation tissue formation following stent placement for major airway stenosis. PATIENTS AND METHODS: Forty-two stent procedures (32 tracheal stents, 3 carinal stents, and 7 bronchial stents) were performed in 40 patients. Seventy-nine stent edge regions (62 tracheal, 17 bronchial stents) were evaluated in this study. RESULTS: Granulation tissue formation was encountered in 11 patients (28.21%). Of the 34 upper ends of evaluated tracheal stents, granulation tissue formation was observed in 6 (17.65%), whereas granulation tissue formation was observed in 2 (7.14%) of the 28 lower ends of tracheal stents evaluated. Of the 17 bronchial stent edge regions, granulation tissue formation occurred in 3 (17.65%) ( P = 0.4352). The rate of granulation tissue formation was higher in those patients with a stent-to-airway diameter ratio of > 90% ( P < 0.0001). Receiver operating characteristic curve analysis further demonstrated that a cut-off stent-to-airway diameter ratio of 90% was effective in predicting granulation tissue formation (AUC: 0.897, Std. error = 0.036, P < 0.0001, 95% CI = 0827-0.968, n = 79). CONCLUSIONS: A stent-to-airway diameter ratio of 90% was found to be the critical cut-off point for predicting granulation tissue formation. Therefore, the optimal stent-to-airway diameter ratio should be ascertained before stent placement.

Extra-long Montgomery T tube in major airway stenosis.

BACKGROUND: We evaluated the efficacy and safety of the extra-long Montgomery T tube for the management of major airway obstruction in tertiary care patients in Taiwan. METHOD: Eleven patients with major airway stenosis treated with an extra-long Montgomery T tube between April 2004 and December 2006 were retrospectively reviewed. Five patients had tracheostomy stenosis, two had intubation stenosis, one had traumatic stenosis, one had corrosive stenosis, one had laser burn stenosis and one had tubercular stenosis. All patients underwent three-dimensional airway reconstruction and endoscopic evaluation of airway stenosis. After determining the severity and location of airway stenosis, rigid bronchotherapy and Montgomery T tube placement were performed by rigid bronchoscopy. RESULTS: The overall procedural success rate was 100 per cent. Three (27 per cent) patients were weaned from artificial ventilation, and all patients exhibited improved respiratory and functional status. No major post-operative complications or mortality were observed. At follow up (mean, 21.5 months), the decannulation rate was 27 per cent, and eight (73 per cent) patients had stable T tube ventilation. In four patients, granulation over the end of the T tube was controlled by endoscopic procedures. Three patients with stents above the vocal folds showed aspiration and required further intervention (i.e. one nasogastric feeding tube for nutrient supplement, one feeding jejunostomy and one stent shortening to decrease aspiration). CONCLUSION: The extra-long Montgomery T tube is an effective and safe method for treating major airway obstruction in the supra-glottic to lower tracheal region.

Salvage surgery after failed chemoradiotherapy in squamous cell carcinoma of the esophagus.

AIMS: To investigate the survival benefit and preoperative risk factors for hospital mortality of salvage surgery in esophageal cancer patients who had locoregional residual/recurrent tumor after definitive chemoradiotherapy. METHODS: We retrospectively reviewed the esophageal cancer patients who presented at our hospital from 1997 to 2004. Forty-seven patients who had squamous cell cancer and developed locoregional recurrent/persistent disease after primary definitive chemoradiotherapy were elected. Twenty-seven of them received salvage esophagectomy (group 1) and the other 20 underwent non-operative treatment only (group 2). In order to assess the surgery-related mobility and mortality in group 1, 191 patients who received neoadjuvant chemoradiotherapy followed by operation during the same time period were also enrolled (group 3). RESULTS: The 5-year overall survival of group 1 patients was 25.4%. In contrast, all of the patients in the group 2 died within 16.7 months. The difference was statistically significant (p=0.0029). In comparison with group 3, group 1 patients had significantly more surgery-related complications and hospital mortality. In univariate analysis for preoperative risk factors, a low albumin or hemoglobulin level was associated with high hospital mortality in group 1 (p=0.004 and 0.003, respectively). After multivariate analysis, only the low albumin level remained borderline significance. As for disease specific survival after salvage surgery, R0 resection was the only independent prognosticator (p=0.049). CONCLUSION: Salvage surgery provides survival benefit in esophageal cancer patients with locoregional persistent or recurrent disease after primary definitive chemoradiotherapy. Preoperative albumin and hemoglobulin levels are associated with hospital mortality and may aid in selecting suitable patient for salvage surgery.

Childhood serum sickness: a case report.

Childhood serum sickness is a rare allergic disease that follows the administration of a foreign antigenic material, most commonly caused by injecting a protein or haptenic drug. The disease is a type III hypersensitivity reaction mediated by deposits of circulating immune complexes in small vessels, which leads to complement activation and subsequent inflammation. The clinical features are fever, cutaneous eruptions, lymphadenopathy, arthralgias, albuminuria, and nephritis. Serum sickness is an acute self-limited disease. We report a 3-year-old child who presented with fever and a rash; an invasive bacterial infection was strongly suspected. He was therefore given penicillin and gentamicin and responded well. At day 4 after admission, he developed a serum sickness reaction and showed symptoms of arthralgias, generalized edema, purpura, and gross hematuria. The white blood cell count was 12 190/mm3 with 7% eosinophils. Urinalysis revealed red blood cell above 100 per high power field, white blood cell 10 to 15 per high power field, and proteinuria. The antibiotics were discontinued and hydrocortisone (20 mg/kg/d), diphenhydramine HCl (4 mg/kg/d), aspirin (66 mg/kg/d) was administered, plus 1 dose of epinephrine (0.01 mL/kg) administered intramuscularly. On day 7, the 3rd day after withholding antibiotics, his condition dramatically improved. The clinical symptoms resolved progressively and his urinalysis returned to normal.

Comparison of antileukemic immunity against U937 cells in atopic asthmatics versus healthy controls.

To assess the antitumor effects in atopic asthmatics versus healthy adults, we designed this study using in vitro mononuclear cells (MNC) culture as an immunity model with human leukemic U937 cells as the target. MNCs were collected from asthmatic subjects and healthy controls. Conditioned media from the MNC cultures (MNC-CM) were collected after stimulation with various concentrations of phytohemagglutinin (PHA). We treated U937 cells with these MNC-CMs, then assayed their proliferation and differentiation after 5 days of culture. At lower PHA doses (1.25 microg/ml), as well as in absence of PHA, the asthmatic MNC-CMs inhibited U937 cells growth to a slightly greater extent than did the MNC-CMs from controls. In contrast, when higher doses of PHA were used (5, 10 microg/ml), this growth-inhibiting effect was dramatically reversed. The dual effect of MNC-CM in these two groups was also shown in U937 cell differentiation assay, assessed as follows: morphological change by Liu's staining, functional change by NBT reduction test and CD 14 expression by flow cytometric detection. We suggest that the antileukemic effects of MNCs from asthmatic patients result from a slightly immunopotentiated status. This immunity may be dramatically reversed, however, after marked activation of MNCs.

Characterization of gammaS-crystallin isoforms from a catfish: evolutionary comparison of various gamma-, gammaS-, and beta-crystallins.

gammaS-Crystallin from catfish eye lenses, formerly designated betas-crystallin in mammalian lenses, is structurally characterized in this study by cDNA cloning and sequencing. To facilitate sequence characterization of gammaS-crystallin with structural properties lying between beta- and gamma-crystallins, a cDNA mixture was constructed from the poly(A)+ mRNA isolated from catfish eye lenses, and amplification by polymerase chain reaction (PCR) was carried out to obtain nucleotide segments encoding multiple gammaS-crystallin isoforms. Sequencing several positive clones revealed that at least two distinct isoforms exist in the gammaS-crystallin class of this teleostean fish, similar to the authentic gamma-crystallin family characterized previously in species of the piscine class. Comparison of protein sequences encoded by two representative catfish gammaS1 and gammaS2 cDNAs with the published sequences of beta-, gamma-, and gammaS-crystallins from shark, carp, bullfrog, bovine, and human lenses indicates that there is about 20-50% sequence homology between catfish gammaS-crystallins and various members of the related beta/gamma-crystallin superfamily from different evolutionary classes, with a higher sequence similarity being found between catfish gammaS- and mammalian gamma-crystallins than between catfish gammaS- and bovine or carp gammaS-crystallins. Phylogenetic trees constructed on the basis of the nucleotide and protein sequence divergence among various beta-, gamma-, and gammaS-crystallins corroborate the closer relatedness of catfish gammaS- to authentic gamma-crystallin than to bovine and carp gammaS-crystallins. The results suggest that evolution of catfish gammaS-crystallins follows a different path from that of bovine and carp gammaS-crystallins and may represent a more ancient offshoot from the ancestral gamma/gammaS coding gene than carp and bovine gammaS-crystallins.

Characterization of gamma-crystallins from a hybrid teleostean fish: multiplicity of isoforms as revealed by cDNA sequence analysis.

gamma-Crystallins were isolated and characterized from the eye lenses of a hybrid species belonging to the teleostean fish. Isoelectric focusing of gamma-crystallin fraction obtained from gel-permeation chromatography revealed that it consists of multiple charge isomers of a protein species with a molecular mass of about 20 kDa. To facilitate the cloning of gamma-crystallin gene, cDNA was constructed from the poly(A)+mRNA isolated from fresh lenses, and amplification by polymerase chain reaction (PCR) was carried out to obtain cDNA encoding multiple gamma-crystallins. Sequencing five of more than 10 positive clones revealed that a multiplicity of isoforms exists in the gamma-crystallin class of teleostean lenses. Comparison of protein sequences encoded by these multiple cDNAs with those published sequences of gamma-crystallins from bovine, mouse an carp lenses indicated that there is about 70-80% sequence homology between different species of piscine species whereas only 50-60% is found between mammals and fishes. Structural analysis of these gamma-crystallins with high methionine contents (11-16%) suggests that there are two major subclasses of piscine gamma-crystallins, i.e. gamma M 1 and gamma M 2, existed long before the appearance of mammalian gamma-crystallin with low methionines.