Saul N. Rajak

Conjunctival Transcriptome in Scarring Trachoma

ABSTRACT Trachoma is a poorly understood immunofibrogenic disease process, initiated by Chlamydia trachomatis. Differences in conjunctival gene expression profiles between Ethiopians with trachomatous trichiasis (with [TTI] or without [TT] inflammation) and controls (C) were investigated to identify relevant host responses. Tarsal conjunctival swab samples were collected for RNA isolation and C. trachomatis PCR. Transcriptome-wide microarray experiments were conducted on 42 samples (TTI, n = 13; TT, n = 15; C, n =14). Specific results were confirmed by using multiplex quantitative reverse transcription-PCR for 16 mRNA targets in an independent collection of case-control samples: 386 case-control pairs (TTI, n = 244; TT, n = 142; C, n = 386). The gene expression profiles of cases were consistent with squamous metaplasia (keratins, SPRR), proinflammatory cytokine production (IL1β, CXCL5, and S100A7), and tissue remodeling (MMP7, MMP9, MMP12, and HAS3). There was no difference in the level of IFNγ between cases and controls. However, cases had increased INDO, NOS2A, and IL13RA2 and reduced IL13. C. trachomatis was detected in 1/772. Cases show evidence of ongoing inflammation and tissue remodeling, which were more marked where clinical inflammation was also present. Significantly, these processes appear to be active in the absence of current C. trachomatis infection. There was limited evidence of a TH1 response (INDO and NOS2A) and no association between a TH2 response and cases. The epithelium appears to be actively involved in late cicatricial stages of trachoma through the production of proinflammatory factors (IL1β, CXCL5, and S100A7). Longitudinal studies are needed to investigate which etiological factors and pathways are associated with progressive scarring and whether simply controlling chlamydial infection will halt progression in people with established cicatricial disease.

Trachomatous Trichiasis and its Management in Endemic Countries

Trichiasis is the sight-threatening consequence of conjunctival scarring in trachoma, the most common infectious cause of blindness worldwide. Trachomatous trichiasis is the result of multiple infections from childhood with Chlamydia trachomatis, which causes recurrent chronic inflammation in the tarsal conjunctiva. This produces conjunctival scarring, entropion, trichiasis, and ultimately blinding corneal opacification. The disease causes painful, usually irreversible sight loss. Over eight million people have trachomatous trichiasis, mostly those living in poor rural communities in 57 endemic countries. The global cost is estimated at US

Conjunctival Expression of Matrix Metalloproteinase and Proinflammatory Cytokine Genes after Trichiasis Surgery

5.3 billion. The WHO recommends surgery as part of the SAFE strategy for controlling the disease.We examine the principles of clinical management, treatment options, and the challenging issues of providing the quantity and quality of surgery that is needed in resource-poor settings.

Pathogenesis of progressive scarring trachoma in Ethiopia and Tanzania and its implications for disease control: two cohort studies.

PURPOSE. Trachoma, the leading infectious cause of blindness, is a chronic inflammatory scarring condition. Blindness follows the development of trichiasis, which is treated surgically. Unfortunately, it frequently recurs, compromising the treatment. In this study, gene expression analysis was used to examine factors that may be involved in the inflammation and tissue remodeling after surgery. METHODS. Subjects were examined before and at 1 and 4 years after surgery. Conjunctival swab samples were collected for bacterial culture, Chlamydia trachomatis PCR, and RNA isolation at 1 year. Quantitative real-time PCR was performed to measure the expression of tumor necrosis factor-alpha (TNF), interleukin-1beta (IL1B), matrix metalloproteinase-1 (MMP1), MMP-2, MMP-9, tissue inhibitor of matrix metalloproteinase 1 (TIMP-1), TIMP-2, and hypoxanthine phosphoribosyl transferase-1 (HPRT1). RESULTS. Two hundred forty individuals with trachomatous trichiasis were recruited. One year after surgery, recurrent trichiasis was associated with a reduced MMP-1/TIMP-1 ratio (P = 0.029). IL1B expression was elevated in the presence of either conjunctival bacterial infection (P = 0.011) or inflammation (P = 0.002). TNF expression was greater in the Mandinka ethnic group (P < 0.0001), and it was increased when clinical inflammation was associated with nonchlamydial bacterial infection (P = 0.012). MMP-9 expression increased when conjunctival inflammation was associated with bacterial infection (P = 0.007). CONCLUSIONS. Recurrent trichiasis was associated with a reduced MMP-1 to TIMP-1 ratio, which may favor the accumulation of fibrotic tissue. Nonchlamydial bacterial infection may induce factors that contribute to conjunctival tissue remodeling and recurrent trichiasis in trachoma. Prospective studies are needed to assess the potential importance of these and other factors in progressive disease.

Trichiasis Surgery in The Gambia: A 4-Year Prospective Study

Background Trachoma causes blindness through a conjunctival scarring process initiated by ocular Chlamydia trachomatis infection; however, the rates, drivers and pathophysiological determinants are poorly understood. We investigated progressive scarring and its relationship to conjunctival infection, inflammation and transcript levels of cytokines and fibrogenic factors. Methodology/Principal Findings We recruited two cohorts, one each in Ethiopia and Tanzania, of individuals with established trachomatous conjunctival scarring. They were followed six-monthly for two years, with clinical examinations and conjunctival swab sample collection. Progressive scarring cases were identified by comparing baseline and two-year photographs, and compared to individuals without progression. Samples were tested for C. trachomatis by PCR and transcript levels of S100A7, IL1B, IL13, IL17A, CXCL5, CTGF, SPARCL1, CEACAM5, MMP7, MMP9 and CD83 were estimated by quantitative RT-PCR. Progressive scarring was found in 135/585 (23.1%) of Ethiopian participants and 173/577 (30.0%) of Tanzanian participants. There was a strong relationship between progressive scarring and increasing inflammatory episodes (Ethiopia: OR 5.93, 95%CI 3.31–10.6, p<0.0001. Tanzania: OR 5.76, 95%CI 2.60–12.7, p<0.0001). No episodes of C. trachomatis infection were detected in the Ethiopian cohort and only 5 episodes in the Tanzanian cohort. Clinical inflammation, but not scarring progression, was associated with increased expression of S100A7, IL1B, IL17A, CXCL5, CTGF, CEACAM5, MMP7, CD83 and reduced SPARCL1. Conclusions/Significance Scarring progressed in the absence of detectable C. trachomatis, which raises uncertainty about the primary drivers of late-stage trachoma. Chronic conjunctival inflammation appears to be central and is associated with enriched expression of pro-inflammatory factors and altered expression of extracellular matrix regulators. Host determinants of scarring progression appear more complex and subtle than the features of inflammation. Overall this indicates a potential role for anti-inflammatory interventions to interrupt progression and the need for trichiasis disease surveillance and surgery long after chlamydial infection has been controlled at community level.

Absorbable Versus Silk Sutures for Surgical Treatment of Trachomatous Trichiasis in Ethiopia: A Randomised Controlled Trial

PURPOSE Trachoma is the leading infectious cause of blindness. Conjunctival Chlamydia trachomatis infection causes scarring, entropion, trichiasis, and blinding corneal opacification. Worldwide, there are 8 million people with trichiasis. Although trichiasis surgery can reduce the risk of blindness, retrospective data suggest that long-term recurrence rates may be high. A 4-year prospective investigation of recurrent trichiasis was conducted in The Gambia. METHODS Patients with trichiasis were examined at baseline, 6 months, 1 year, and 4 years after posterior lamellar tarsal rotation surgery. Conjunctival swabs for bacteriology and PCR for C. trachomatis were collected at baseline, 6 months, and 1 year. RESULTS Three hundred fifty-six Gambian patients were enrolled at baseline and 266 were reassessed at 4 years (94% of surviving patients). The recurrence rates were 32%, 40%, and 41% at 6 months, 1 year, and 4 years, respectively. At 4 years, 30% of patients had bilateral trichiasis and 21% had bilateral corneal opacity. Recurrence was associated with severe conjunctival inflammation and severe trichiasis (>10 lashes) at baseline. CONCLUSIONS Trichiasis recurrence rates were high, and most cases recurred within 6 months of surgery. The results suggest that there are important aspects of surgical technique and quality that should to be addressed. Persistent inflammation is strongly associated with recurrence at 4 years.

Surgery Versus Epilation for the Treatment of Minor Trichiasis in Ethiopia: A Randomised Controlled Noninferiority Trial

In this randomized trial, Saul Rajak et al. compare silk sutures (removed at 7–10 days) or absorbable sutures (left in place) during surgery for the management of trachomatous trichiasis.

Clearing the Backlog: Trichiasis Surgeon Retention and Productivity in Northern Ethiopia

In this randomized, non-inferiority trial, Saul Rajak et al compare epilation and surgery for the management of minor trichiasis in Ethiopia, the country with the most cases of trachomatous trichiasis.

The clinical phenotype of trachomatous trichiasis in Ethiopia: not all trichiasis is due to entropion.

Background In 2006 there were an estimated 645,000 people in Amhara, Ethiopia, with trachomatous trichiasis (TT) who needed surgery. Despite an extensive integrated eye care worker training programme (IECW) and robust support for TT surgical services, productivity has not reached targets. We investigated why surgeon productivity was below target. Methodology/Principal Findings Confidential interviews were conducted in person with TT surgeons trained from 24 selected districts in Amhara Region and their supervisors. Determinants of attrition and productivity were investigated. We interviewed 225 people who had received IECW training; 139 (59%) had subsequently changed career/job. Staff retention was associated with good road access to their health centre, mobile telephone network and a shorter time from initial training. Amongst the 94 IECW still working in the programme, the average number of patients operated was 41/year, which was mostly (86%) done through outreach campaigns and only 14% of cases were performed in the static facilities where they routinely worked. Spot checks were made of surgical instruments and consumables: only 3/94 IECW had the minimum instruments and consumables to perform surgery. The main barriers to operating were lack of time, shortage of consumables, lack of patients, lack of support and equipment problems. Very few IECW received ongoing supervision or active management. Conclusions/Significance Surgeon attrition rates are high. Vertical surgery campaigns were effective in treating large numbers of cases, whilst static-site service productivity was low. Good health system management is key to building a well-staffed and well-run service.

Posterior lamellar versus bilamellar tarsal rotation surgery for trachomatous trichiasis in Ethiopia: a randomised controlled trial

PURPOSE Trachomatous trichiasis (TT) is usually described as a cicatricial entropion of the upper lid; however, other forms of trichiasis have been reported. This variation in clinical phenotype is potentially important for treatment guidelines. Therefore, this study was conducted to investigate the range of disease type and severity encompassed by TT. METHODS Individuals presenting with TT to surgical treatment campaigns were examined by a single ophthalmologist using the Detailed WHO Trachoma Grading System. Additional features were graded, including type of trichiatic lashes (metaplastic, misdirected, and entropic), lower lid trichiasis, entropion severity, and lid margin mucocutaneous junction (MCJ) position. RESULTS Recruited were 2556 individuals with previously unoperated TT in at least one eye (4310 eyes). The median number of lashes touching the eye was 2 (range, 0 [epilating]-133). Entropion was absent or mild in 2328 (54.0%) eyes, moderate in 1259 (29.2%) eyes, and severe in 723 (16.8%) eyes. Trichiatic lashes were predominantly metaplastic or misdirected (80.2%), rather than secondary to entropion; 4204 (97.7%) had anteroplacement of the MCJ; and lower lid trichiasis was present in 494 (11.5%). Entropion was more severe among those with a low BMI, those who were female, those aged less than 50 years, and those with moderate to severe conjunctival inflammation, central corneal opacity, and severe conjunctival scarring. CONCLUSIONS Many patients with TT have minimal or no entropion. The trichiasis is frequently attributable to metaplastic or misdirected eyelashes. The results of tarsal rotation surgery in TT patients without manifest entropion should be investigated and potentially alternative treatment strategies evaluated.