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The human immunodeficiency virus type 1 (HIV-1)-encoded RNA-binding protein Tat is known to play an essential role in viral gene expression. In the search for novel compounds to inhibit Tat transactivity, one coumarin derivative, BPRHIV001, was identified, with a 50% effective concentration (EC(50)) against HIV-1 at 1.3 nM. BPRHIV001 is likely to exert its effects at the stage after initiation of RNAPII elongation since Tat protein expression and the assembly of the Tat/P-TEFb complex remained unchanged. Next, a reduction of the p300 protein level, known to modulate Tat function through acetylation, was observed upon BPRHIV001 treatment, while the p300 mRNA level was unaffected. A concordant reduction of phosphorylated Akt, which was shown to be closely related to p300 stability, was observed in the presence of BPRHIV001 and was accompanied by a decrease of phosphorylated PDPK1, a well-known Akt activator. Furthermore, the docking analysis revealed that the reduced PDPK1 phosphorylation likely resulted from the allosteric effect of interaction between BPRHIV001 and PDPK1. With strong synergistic effects with current reverse transcriptase inhibitors, BPRHIV001 has the potential to become a promising lead compound for the development of a novel therapeutic agent against HIV-1 infection.

Background: Post-cardiopulmonary bypass (CPB) lung injury is the combination of whole body inflammatory response and local ischemia-reperfusion (IR) injury. We investigated the benefit of pulmonary perfusion with L-arginine in protection against post-CPB lung injury., Methods: New Zealand white rabbits (n = 50, weight, 2.5-2.8 kg) were divided into five groups (n = 10 each): sham (sham sternotomy), CPB (CPB without pulmonary perfusion), perfusion (CPB with pulmonary perfusion), L-arginine (CPB with perfusion + L-arginine), and L-NAME (CPB with perfusion + L-NAME). The duration of CPB was 60 min followed by 2 h of reperfusion. Pulmonary perfusion was performed every 20 min through the pulmonary artery during CPB. Checking parameters included: (1) pulmonary vascular resistance, (2) pulmonary artery endothelium relaxation (organ chamber study), and (3) IR marker (myeloperoxidase) and inflammatory markers (TNF-α, IL-B, NF-κB)., Results: CPB induced pulmonary artery endothelium dysfunction manifested by increased pulmonary vascular resistance and impaired pulmonary artery relaxation. Pulmonary perfusion could significantly reverse the phenomenon (P < 0.01) while provision of NO precursor-L-arginine with pulmonary perfusion together further possessed significant relaxation ability for pulmonary arterial endothelium compared with perfusion alone (P < 0.05). Accordingly, lung parenchyma myeloperoxidase activity and inflammatory cytokine level were also markedly increased after CPB (P < 0.05). Pulmonary perfusion could partially decrease the response, whereas additional L-arginine further attenuated inflammatory cytokine release (P < 0.05)., Conclusions: Pulmonary perfusion during CPB partially ameliorates CPB-induced lung injury. Pulmonary perfusion with L-arginine could further attenuate lung injury by restoring endothelial function and decreasing inflammatory response., (Copyright © 2011 Elsevier Inc. All rights reserved.)

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., (© Georg Thieme Verlag KG Stuttgart · New York.)

Background: Close circumferential resection margin (CRM) is an established predictor for locoregional recurrence (LR) in rectal cancer but remains controversial in esophageal malignancy. As yet, little is known about the significance of CRM after chemoradiotherapy (CRT), especially in squamous cell carcinoma (SCC). This study investigated the relationship between CRM distance and recurrence after neoadjuvant CRT in esophageal SCC patients., Methods: Between 1997 and 2005, esophageal SCC patients who underwent surgery after neoadjuvant CRT and with pathology stage T3N0M0 and T3N1M0 (metastatic lymph nodes <2) were selected. CRM distance was reassessed and divided into three groups (group 1: CRM >1 mm, group 2: uninvolved CRM but <1 mm, group 3: CRM involved)., Results: The cohort comprised 145 male and 6 female patients with mean age of 57 years. There were 74, 51, and 26 patients in group 1, 2, and 3, respectively. With median follow-up period of 50 months, LR developed in 30.5% of patients. Both group 2 and group 3 had significantly higher LR than group 1 (37 and 42% vs. 21%, P < 0.05). Meanwhile, mean time from operation to recurrence was also significantly shorter in group 2 and group 3 than in group 1 (267 and 269 days versus 402 days, P < 0.05). Five-year disease-specific survival (DSS) was highest in group 1 (40%). Despite the similarity in LR, 5-year DSS significantly differed between group 2 and group 3 (22 vs. 7%, P < 0.05). The higher rate of distant recurrence (DR) and concomitant LR + DR in group 3 accounted for the survival difference., Conclusion: In ypT3 esophageal SCC patients, CRM distance provides useful information for risk stratification in cancer recurrence and survival.

Cell signaling pathways play important roles in oncogenesis. Among a large number of signaling regulators in different pathways, 4E-binding protein 1 (4E-BP1) was found to be a key factor, which converges several oncogenic signals, phosphorylates the molecules, and drives the downstream proliferative signals. Recent studies showed that high expression of phosphorylated 4E-BP-1 (p-4E-BP1) is associated with poor prognosis, tumor progression, or nodal metastasis in different human cancers, but its prognostic significance in esophageal cancer remains undefined. In this study, we investigated the expression levels of p-4E-BP1 with two different phosphorylation sites Thr(37/46) and Thr(70) by immunohistochemistry and their prognostic significance in 78 cases of surgically resected esophageal squamous cell carcinoma (SCC) for the first time. We found no correlation of p-4E-BP1 expression with age, gender, preoperative concurrent chemoradiotherapy, tumor grade, pT classification, pN, pM, or pStage. Multivariate Cox regression analysis showed that high expression of p-4E-BP-1 Thr(37/46) was an independent adverse prognostic factor, with a hazard ratio of 1.73 (95% confidence interval = 1.03-2.90) and a p value of 0.038. Stratifying the patients with other prognostic factors, we found that the effect of p-4E-BP1 Thr(37/46) on survival was significant only in patients with relatively early stage disease (pT1/pT2, pN0, or pStage I/II; p = 0.0047, 0.012, and 0.011, respectively). Our data suggest that assessment of p-4E-BP1 expression could identify a subpopulation of earlier stage esophageal SCC patients with poor prognosis. These patients could be possible candidates for future studies on more aggressive treatment or target therapy.

Background: Non-regional lymph node metastasis in intrathoracic esophageal cancer is classified as M1 lesion with poor prognosis following surgery alone. We studied the controversial question of whether chemoradiotherapy (CRT) improves survival of these patients., Methods: A cohort of patients with clinically overt nodal M1 disease, which could be encompassed by a tolerable radiation therapy port, was selected from the database of the Chang Gung Memorial Hospital., Results: From 1994 to 2005, 54 nodal stage IV intrathoracic esophageal squamous cell carcinoma (SCC) patients received neoadjuvant CRT. Significant response occurred in 24 patients. Scheduled esophagectomy was performed in 26 patients. The 3-year overall survival (OS) and disease-free survival (DFS) for the whole group were 27% (median: 14.2 months) and 22% (median: 14.7 months), respectively. Multivariate analysis identified pretherapy lymph nodes classified as M1a and R0 resection after CRT as independent favorable prognosticators. Median survival reached 36.9 months in the pretherapy M1a subgroup as opposed to 12.5 months in the M1b subgroup (3-year-DFS: 40% vs. 10%, P = 0.0117). Scheduled surgery after CRT benefits only after R0 resection (3-year-DFS: 36%, median survival: 45 months). The group with incomplete resection had a high surgical risk and dismal survival compared to the non-surgery group (3-year-DFS: 0% vs. 9%, 9.5 vs. 10.5 months)., Conclusions: Pretherapy M1a disease had a significantly better survival than nodal M1b disease after CRT in SCC. Aggressive surgical treatment after CRT is reserved for cases when complete resection is anticipated., ((c) 2010 Wiley-Liss, Inc.)

Background: Pathological complete response (pCR) after chemoradiotherapy (CRT) is the best predictor of survival in patients with squamous cell carcinoma (SCC) of the esophagus. Although no adjuvant treatment is recommended for individuals who achieve pCR, approximately 30% of these patients develop a recurrence. We applied an immunohistochemistry (IHC) stain on resected lymph nodes to evaluate the incidence of lymph node micrometastases (LNMs) and its prognostic significance., Methods: Between 1995 and 2004, paraffin-embedded specimens for evaluating the presence of LNM were available for 52 pCR patients. We performed IHC staining with the cytokeratin antibody AE1/AE3 on resected lymph nodes and correlated the pathological findings to cancer recurrence and survival., Results: A total of 510 lymph nodes from 52 patients were examined. The incidence of LNM in pathological complete responders after CRT was 11.5%. Only six patients were found to have AE1/AE3-positive cells in the IHC staining. However, patients with LNM had a higher recurrence rate (50% vs. 22%, P = 0.16) and a reduced disease-free survival rate (44% vs. 75%, P = 0.16). Patients with LNM had a 3.6-fold higher probability to develop recurrence compared with those without LNM (P = 0.15)., Conclusion: LNMs are uncommon in pathological complete responders but hold prognostic significance.

Podoplanin is a mucin-like glycoprotein expressed in the lymphatic endothelium. It has been suggested to play a role in lymphangiogenesis, since podoplanin deficient mice were found to have dilated malfunctioning lymphatic vessels and lymphedema. High podoplanin expression in tumor cells was found to correlate with lymph node metastasis and poor clinical outcome in patients with oral squamous cell carcinoma (SCC). However, the prognostic significance of podoplanin expression in esophageal SCC remains unexplored. Herein, we studied podoplanin expression in 59 patients who underwent surgical resection of esophageal SCC, with 43 of them preceded by preoperative concurrent chemoradiotherapy (CCRT). We found that high podoplanin expression strongly correlated with clinical nodal metastasis (cN1; p=0.0063), which was associated with short survival (p=0.012). However, there was no direct association between high podoplanin expression and short survival. We also found that lymphatic vessel invasion in the resected esophagus was strongly associated with pathological nodal metastasis (pN1; p=0.00092). Our results suggest that podoplanin could also play a role in tumor aggressiveness in esophageal SCC, as well as in oral SCC.

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.

Background: The benefit of bullae ablation for the management of primary spontaneous pneumothorax (PSP) has been established. However, various modalities for bullae ablation have been reported from different centers., Objective: The present study aimed to assess whether endoloop ligation of bullae was as effective as staple bullectomy for preventing the recurrence of pneumothorax., Methods: Between January 1993 and December 2003, 226 patients (203 men and 23 women) with PSP were recorded and retrospectively reviewed. One hundred thirty (57.5%) patients were treated with endoloop ligation of bullae and the other 96 (42.5%) were treated with staple bullectomy. Mechanical abrasion was performed in all patients after bullae ablation., Results: The recurrence rate of pneumothorax was 6.2% (8 patients) in the endoloop ligation group and 17.7% (17 patients) in the staple bullectomy group (p = 0.006). The postoperative complication rate was 14.6 and 20.8% in the endoloop ligation and staple bullectomy groups, respectively (p = 0.221). The long-term chest discomfort rate after endoloop ligation and staple bullectomy was 14.6 and 13.5%, respectively (p = 0.819), Conclusions: Endoloop ligation of bullae is as effective as mechanical staple bullectomy for the management of bullae in primary spontaneous pneumothorax.

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.

Background: Preoperative chemoradiotherapy (CRT) followed by esophagectomy is becoming one of the standard treatment strategies for esophageal cancer. Pathologic complete response (pCR) after CRT is the best predictor of survival in squamous cell carcinoma (SCC) of the esophagus. Although no adjuvant treatment is recommended for individuals who achieve pCR, approximately 30% of these patients develop recurrence. Herein we sought to retrospectively investigate the independent predictors of tumor recurrence in this patient group., Methods: Between 1995 and 2004, we investigated seventy patients (69 males and 1 female; mean age: 56.1 years) with esophageal SCC who achieved pCR following preoperative chemoradiotherapy. Study end points included tumor recurrence, disease-specific survival (DSS), and disease-free survival (DFS). Univariate and multivariate analyses were used to identify risk factors for the study end points., Results: Mean follow-up time for patients who survived was 65.8 months. At the time of analysis, 18 patients (25.7%) died of the disease and 22 patients (31.4%) developed recurrence. Multivariate analysis showed that pretherapy T3-4 disease was the most important adverse factor for tumor recurrence (P = 0.007), DFS (P = 0.005), and DSS (P = 0.026). The 5-year DFS was 45% for patients with clinical T3-4 disease and 85% for those with clinical T1-2 disease (P = 0.02)., Conclusions: We have shown that up to 31.4% of patients with esophageal SCC who achieve pCR develop tumor recurrence thereafter. Pretherapy T3-4 disease was a strong and independent adverse risk factor for 5-year tumor recurrence, DSS and DFS. High-risk patients with T3-4 disease should be followed with a strict surveillance protocol.

Background: Although elderly patients with thoracic disease were considered to be poor candidates for thoracotomy before, recent advances in preoperative and postoperative care as well as surgical techniques have improved outcomes of thoracotomies in this patient group. The aim of this study was to investigate surgical risk factors and results in elderly patients (aged > or =70 years) with thoracic empyema., Methods: Seventy-one elderly patients with empyema thoracis were enrolled and evaluated from July 2000 to April 2003. The following characteristics and clinical data were analysed: age, sex, aetiology of empyema, comorbid diseases, preoperative conditions, postoperative days of intubation, length of hospital stay after surgery, complications and mortality., Results: Surgical intervention, including total pneumonolysis and evacuation of the pleura empyema cavity, was carried out in all patients. Possible influent risk factors on the outcome were analysed. The sample group included 54 men and 17 women with an average age of 76.8 years. The causes of empyema included parapneumonic effusion (n = 43), lung abscess (n = 8), necrotizing pneumonitis (n = 8), malignancy (n = 5), cirrhosis (n = 2), oesophageal perforation (n = 2), post-traumatic empyema (n = 2) and post-thoracotomy complication (n = 1). The 30-day mortality rate was 11.3% and the in-hospital mortality rate was 18.3% (13 of 71). Mean follow up was 9.4 months and mean duration of postoperative hospitalization was 35.8 days. Analysis of risk factors showed that patients with necrotizing pneumonitis or abscess had the highest mortality rate (10 of 18, 62.6%). The second highest risk factor was preoperative intubation or ventilator-dependency (8 of 18, 44.4%)., Conclusion: This study presents the clinical features and outcomes of 71 elderly patients with empyema thoracis who underwent surgical treatment. The 30-day surgical mortality rate was 11.3%. Significant risk factors in elderly patients with empyema thoracis were necrotizing pneumonitis, abscess and preoperative intubation/ventilation. This study also suggested that surgical treatment of empyema thoracic in elderly patients is recommended after failed conservative treatment because of the acceptably postoperative complication and mortality rate.

Stenting is an effective method of treating airway stenosis. However, airway obstruction due to mucus plugging is problematic. Treatment of this has mainly been conservative. This report describes a patient with complicated tracheal stenosis treated with silicone hood stent, who experienced repeated episodes of airway obstruction due to sputum accumulation. A novel method of sputum clearance and self-care, using a long endotracheal tube within the silicone stent, is described. This procedure resulted in a stable and patent airway, and the patient was discharged smoothly after the procedure. No further respiratory symptom occurred during the follow-up period.

Managing the airway of patients with critical tracheal stenosis remains a formidable challenge to surgeons and anesthiologists. Various methods for controlling the airway have been established to solve this problem. This study describes the critical tracheal stenosis is managed by dilating and coring technique under direct vision with a rigid bronchoscope. From 2001 to 2003, 34 patients (23 males/11 females) with critical tracheal stenosis (>90%) underwent 37 surgical interventions. The etiologies included 8 cases of post-intubation, 7 cases of post-tracheostomy, 5 cases of post-anastomotic, 14 cases of endotracheal tumor (4 primary, 10 secondary), and 3 cases (1 tuberculosis, 1 laser burn, and 1 idiopathic) of other etiologies. All procedures could achieve airway control with rigid bronchoscope. Other endoscopic procedures (laser, and stent placement) were required to achieve a patent airway and seven patients received airway reconstruction as a definite procedure. Rigid bronchoscopy could provide good results for controlling airway in patients with critical trachea stenosis.

Purpose: The high mortality associated with esophageal perforation can be reduced by aggressive surgery and good critical care. We report our experience of treating esophageal perforation in a clinic in Taiwan., Methods: The subjects were 28 patients who underwent surgery for a benign esophageal perforation., Results: The esophageal perforation was iatrogenic in 11 patients, spontaneous in 8, and caused by foreign body injury in 9. Most (22/28) of the patients were seen longer than 24 h after perforation, and 77% had empyema preoperatively. The perforation was located in the cervical area in 5 patients and in the thoracic esophagus in 23. We performed primary repair in 24 patients, esophagectomy in 3, and drainage in 1. Leakage occurred after primary repair in ten (41%) patients, resulting in one death, and two patients died of other diseases. Postoperative leakage prolonged the hospital stay but had no impact on mortality. Overall survival was 90%. Univariate analysis revealed that age, timing of treatment, and cause and location of the perforation influenced outcome, but multivariate analysis failed to identify a predictor of mortality., Conclusions: Early diagnosis and intervention are crucial to prevent morbidity and mortality in patients with esophageal perforation. Primary repair is feasible even if the diagnosis is delayed.

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.

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.

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., (Copyright 1998 Academic Press.)

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.