Liver Transplantation: A Test of Cellular Physiology, Preservation, and Injury.

Liver transplantation has evolved into a mature clinical field, but scarcity of usable organs poses a unique challenge. Expanding the donor pool requires novel approaches for protecting hepatic physiology and cellular homeostasis. Here we define hepatocellular injury during transplantation, with an emphasis on modifiable cell death pathways as future therapeutics.

Tattoo complications in treated and non-treated psoriatic patients.

BACKGROUND: Tattooing is a widespread phenomenon, with an estimated prevalence of 10-30% in Western populations. For psoriasis patients, current recommendations are to avoid having a tattoo if the disease is active and they are receiving immunosuppressive treatments. Although scientific data supporting these recommendations are lacking, dermatologists are often reluctant to advocate tattooing in psoriasis patients. OBJECTIVE: We aimed to evaluate the frequency of tattoo complications in patients with psoriasis and determine whether the occurrence of complications was associated with psoriasis status and treatments received at the time of tattooing. METHODS: We performed a multicentre cross-sectional study. Adults with psoriasis were consecutively included and classified as tattooed or non-tattooed. Prevalence of complications associated with tattoos was then evaluated according to psoriasis onset and treatments. The study was divided into three parts, in which data were collected through a series of questionnaires filled in by the dermatologist. Complications included pruritus, oedema, allergic reaction/eczema, infection/superinfection, granuloma, lichenification, photosensitivity, Koebner phenomenon and psoriasis flare after tattooing. Diagnosis of complications was made retrospectively. RESULTS: We included 2053 psoriatic patients, 20.2% had 894 tattoos. Amongst non-tattooed patients, 15.4% had wished to be tattooed, with psoriasis being stated as a reason for not having a tattoo by 44.0% and 5.7% indicating that they planned to have a tattoo in the future. Local complications, such as oedema, pruritus, allergy and Koebner phenomenon, were reported in tattoos in 6.6%, most frequently in patients with psoriasis requiring treatment at the time of tattooing (P < 0.0001). No severe complications were reported. CONCLUSIONS: The rate of tattoo complications in psoriasis patients was low. Although the risk of complications was highest amongst patients with psoriasis requiring treatment at the time of tattooing, all the complications observed were benign. These results can be helpful for practitioners to give objective information to patients.

[Leg ulcers occurring under tyrosine kinase inhibitor therapy (sunitinib, nilotinib)]. / Ulcères des membres inférieurs développés sous inhibiteurs de tyrosine kinase (sunitinib, nilotinib).

BACKGROUND: Certain anticancer drugs are known to induce leg ulcers, mainly chemotherapy agents such as hydroxyurea. We report 2 cases of leg ulcers in cancer patients treated with the tyrosine kinase inhibitors, sunitinib and nilotinib, and we discuss the role of these treatments in the pathogenesis of leg ulcers. PATIENTS AND METHODS: Case 1. A 62-year-old patient on sunitinib for intrahepatic cholangiocarcinoma developed a lesion on her right foot. The vascular evaluation was negative. After progressive worsening, sunitinib was stopped and healing was observed within a few months. Case 2. A 83-year-old patient had been treated for chronic myeloid leukemia since 2005. Nilotinib was introduced in 2009. Peripheral arterial revascularization was required in May 2013. A few months later, worsening was noted with the onset of ulceration and necrosis of the third toe. Further revascularisation surgery was performed, and nilotinib was suspended and antiplatelets introduced. Healing occurred a few months later. DISCUSSION: Many skin reactions have been described in patients on nilotinib and sunitinib, but few publications report the development of de novo ulcers in patients without risk factors. The pathophysiology of the development of ulcers in patients receiving tyrosine kinase inhibitors is not clear, and probably involves several mechanisms of action. The increasing use of this type of treatment could lead to an upsurge in the incidence of vascular complications. CONCLUSION: We report two cases of leg ulcers developing in patients on tyrosine kinase inhibitors and raise the question of causal implication of these treatments in the pathogenesis of ulcers.

[Subcutaneous calcinosis, a late and severe complication of radiotherapy: Three cases]. / La calcinose sous-cutanée, une complication rare et sévère de la radiothérapie: trois observations.

BACKGROUND: Radiation-induced subcutaneous calcinosis is a rare and special form of potentially severe subcutaneous calcinosis of late onset. Herein, we report three cases of this disease, occurring in each instance more than 10 years after use of radiotherapy as an adjuvant treatment in breast cancer. PATIENTS AND METHODS: Our report concerns 3 women aged 69-88 years consulting for pre-sternal ulcers (n=2) and/or subcutaneous nodules (n=2). These lesions developed on areas irradiated between 10 and 38 years earlier for breast cancer. In all three cases, radiological explorations showed extensive subcutaneous calcification. In one case, calcification extended into the mediastinum. In each patient, a diagnosis of radiation-induced subcutaneous calcinosis was made and symptomatic treatment was given. DISCUSSION: Radiation-induced subcutaneous calcinosis is an irreversible and rare complication of high-dose radiation that usually occurs several years after radiotherapy. Its severity is related to potential ulcerations, pain and a risk for in-depth extension up to the mediastina. This complication remains unclear and treatment has not been codified. The only option seems to be "heavy" plastic surgery.

[Crusted scabies induced by topical corticosteroids: A case report]. / Gale hyperkératosique favorisée par l'utilisation de dermocorticoïdes : à propos d'un cas.

The frequency of scabies is increasing in France. Crusted (or Norwegian) scabies is a very contagious form of scabies because of the huge number of mites in the skin. It is observed in patients suffering from immunodepression, motor or sensory deficiency, or mental retardation. The clinical presentation, except for the classic manifestation of scabies, is characterized by crusted lesions. Treatment is not easy and requires hospitalization. Topical corticosteroids are frequently used for children's dermatological diseases. Their long-term and inappropriate application in an infested scabies child can induce crusted scabies. We report on a case of an 8-year-old boy who developed crusted scabies induced by topical corticosteroid application. We discuss the therapeutic aspects of this severe form of scabies.

[Compression care for venous leg ulcers: Assessment of medical practices in 100 patients]. / La compression dans la prise en charge des ulcères veineux des membres inférieurs. Évaluation des pratiques professionnelles sur 100 patients.

INTRODUCTION: The etiologic treatment of venous ulcers is based on compression therapy in compliance with the new guidelines promulgated by the French National Authority for Health (HAS) in 2010. Prescriptions often originate from a request by the nurse delivering care in the patient's home. A recent French study demonstrated the positive impact of compression therapy on venous ulcer healing. The objective of this study was to evaluate medical practices in order to target corrective actions. MATERIALS AND METHODS: We conducted a single-center prospective observational study, using a standardized questionnaire from January to May 2014. Patients with venous ulcers who had an indication for compression therapy were included consecutively. The questionnaire collected demographic and clinical data and also recorded the results of complementary tests and the characteristics of the compression therapy. RESULTS: One hundred patients were included (61 women and 39 men). The average age was 76 years. Patients were recruited during consultations (n = 69), with a majority of patients living at home (n = 80) and receiving home care delivered by a nurse (n = 81). Thirteen patients were seen for the first time and 87 patients were receiving long-term care. The ulcers evolved for 5.7 years on average. Patients presented peri-lesional edema (n = 58), ankle ankylosis (n = 49), autonomous mobilization (n = 40) and walking problems (n = 60). Physical therapy was prescribed for 39 patients and was effectively carried out for 24. The two main causes were venous varices (n = 66) and post-phlebitis disease (n = 18). Compression therapy was prescribed for 97 patients and the products delivered by the pharmacy were consistent with the prescription for 74 patients. Compliance with compression therapy was faulty for 28 patients because of poor tolerance, misunderstanding, manipulation problems, or inappropriate footwear. At assessment, 66 patients were wearing the bands, but not always correctly (starting at the base of the toes [n = 61], heel included [n = 43], proper stretching [n = 43] up to below the knee [n = 57]). Proper footwear was noted in 70 patients. CONCLUSION: Data are scarce on compliance with compression banding. This study shows that further efforts are needed to ensure proper patient education and professional training for physicians and allied profession concerning the installation of compression therapy. Total compliance was observed in only 35% of patients. In addition, the products delivered by the pharmacy were not consistent with the prescription in 26% of cases. Many discrepancies were observed between what was prescribed and what the patients achieved. Patient adherence is a crucial issue for compression therapy.

[Acute finger ischemia: A retrospective study of 13 patients]. / Ischémie digitale aiguë : étude rétrospective de 13 cas.

BACKGROUND: Acute ischemia of the upper limbs is rare in comparison with ischemia of the lower limbs. The origins of this condition are varied. GOALS: We retrospectively analyzed cases of acute finger ischemia (Raynaud's phenomena was excluded) in a dermatology department between 2008 and 2013 in order to evaluate the etiology and management of this phenomenon. RESULTS: Thirteen cases of finger ischemia were reported. The mean age was 54 years. Active smoking was noted in 11 cases. Ischemia was acute in 9 cases and subacute in 4 cases. The location was unilateral in 10 cases and bilateral in 2. Etiologies were: dysplasia of the palmar arch, antiphospholipid antibody syndrome, frostbite, distal arteritis linked to smoking, paraneoplastic arteritis, Buerger's disease, polyarteritis nodosa, stenosis of the subclavian artery, and 3 cases of embolic origin (ulnar, cardiac, and paraneoplastic aneurysm). In the acute phase, antiplatelets were given in 6 cases, anticoagulants in 10 cases and ilomedin in 6 cases. Sympathectomy was performed in 1 case and amputation in 2 cases. DISCUSSION: This study illustrates the diversity of etiologies of finger ischemia. The etiological test battery should be broad and include immunological and thrombophilia tests, arterial and cardiac investigations, cervical radiography and CT scan (screening for cancer). Close collaboration between dermatologists, hematologists, vascular surgeons and radiologists is essential for the management of these patients.

[Cutaneous tuberculosis and erythema induratum: a retrospective study of 13 cases in France]. / Tuberculose cutanée et érythème induré de Bazin: étude rétrospective de 13 cas.

BACKGROUND: Tuberculosis is the most common mycobacterial disease in the world. The cutaneous form is rare in low endemic countries. The occurrence of several cutaneous tuberculosis cases in our dermatology department during 2011-2012 led us to investigate whether there was a resurgence of cutaneous tuberculosis in France. The aim was to analyse changes in cutaneous tuberculosis and the related clinical, microbiological and therapeutic data. PATIENTS AND METHODS: We conducted a retrospective study in our hospital between 2005 and 2012 by querying the PMSI database (code: A 18.4). Epidemiological, clinical, paraclinical and therapeutic data were collected. Erythema induratum was regarded as a variety of cutaneous tuberculosis. RESULTS: Thirteen patients presented cutaneous tuberculosis between 2005 and 2012. The most frequent clinical forms were erythema induratum of Bazin (n=6) and scrofuloderma (n=3). Microbiological evidence was provided in only 4 cases. DISCUSSION: Diagnosis is difficult due to the varied clinical forms and to the relatively high frequency of paucibacillary forms. Further, the set of additional examinations is non-specific. In some cases, it is only therapeutic tests that allow diagnosis to be made. The place of new diagnostic tools must be clarified and a universally acceptable definition of erythema induratum devised.

[Two cases of severe pruritus revealing Hodgkin's disease in children]. / Prurit sévère révélant un lymphome de Hodgkin chez l'enfant : deux observations.

BACKGROUND: Pruritus in children is a frequent reason for consultation, most often related to a common dermatosis. Where dermatological investigation fails to reveal a dermatological cause, a general cause may be suspected. We report three cases of pruritus revealing Hodgkin's lymphoma in children. PATIENTS AND METHODS: Case 1: a 14-year-old girl presented pruritus with diffuse scratching lesions present for 6 months, associated with right cervical lymph nodes occurring after the onset of pruritus. Tomodensitometry revealed involvement of the supra- and sub-diaphragmatic lymph nodes as well as pulmonary involvement. Lymph node biopsy confirmed nodular sclerosing Hodgkin's lymphoma. Case 2: a 14-year-old boy was hospitalized for suspected psychogenic pruritus. He presented intense itching, predominantly in the lower extremities and at night, occurring over the previous 6 months as well as night sweats. Examination showed that the patient had lost 5kg in 1 month and had a low-grade fever of 38°C; he presented linear striated scratching lesions on both legs. Cervical and inguinal lymphadenopathy was seen. The chest scan also revealed supra-diaphragmatic adenomegalies. The biopsy confirmed Hodgkin's lymphoma. DISCUSSION: Systemic causes of pruritus in children are poorly described in the literature. In these two cases, pruritus allowed a diagnosis of Hodgkin's lymphoma to be made, emphasizing the important role of dermatologists in the early diagnosis of haematological malignancy.

[Phlegmasia cerulea dolens]. / La phlébite bleue.

BACKGROUND: Herein we report a case of phlegmasia cerulea dolens, a form of venous thrombosis complicated by arterial ischaemia. PATIENTS AND METHODS: A 69-year-old man presented a bilateral trophic condition of the lower limbs that had appeared 3 weeks earlier. The patient had a history of metastatic urothelial bladder carcinoma and arteritis. Clinical examination revealed right leg ulcers with massive bilateral oedema of the lower limbs, cyanosis and local ischaemia. Doppler ultrasound revealed bilateral and proximal deep vein thrombosis (sural and superficial femoral veins of the right leg; sural and iliac veins of the left leg) without any distal arterial flow. We concluded on a diagnosis of bilateral phlegmasia cerulea dolens. DISCUSSION: Phlegmasia cerulea dolens is a particular type of deep venous thrombosis in which a proximal venous thrombus is combined with arterial ischaemic signs due to brutal and massive oedema and slowing down of arterial flow. In most cases, the lower limbs are involved, with malignancy being the most common cause. It should be suspected in the presence of the classical triad of "pain, oedema and cyanosis", with confirmation by Doppler ultrasound. There is no general consensus regarding standard management. Traditionally, systemic anticoagulation has been the mainstay of treatment for this condition. Endovascular surgery may be a possibility in some cases. Prompt diagnosis and rapid treatment initiation are paramount in order to improve the prognosis of this severe condition with ominous prospects.

Ca²âº signals promote GLUT4 exocytosis and reduce its endocytosis in muscle cells.

Elevating cytosolic Ca(2+) stimulates glucose uptake in skeletal muscle, but how Ca(2+) affects intracellular traffic of GLUT4 is unknown. In tissue, changes in Ca(2+) leading to contraction preclude analysis of the impact of individual, Ca(2+)-derived signals. In L6 muscle cells stably expressing GLUT4myc, the Ca(2+) ionophore ionomycin raised cytosolic Ca(2+) and caused a gain in cell surface GLUT4myc. Extra- and intracellular Ca(2+) chelators (EGTA, BAPTA-AM) reversed this response. Ionomycin activated calcium calmodulin kinase II (CaMKII), AMPK, and PKCs, but not Akt. Silencing CaMKIIδ or AMPKα1/α2 partly reduced the ionomycin-induced gain in surface GLUT4myc, as did peptidic or small molecule inhibitors of CaMKII (CN21) and AMPK (Compound C). Compared with the conventional isoenzyme PKC inhibitor Gö6976, the conventional plus novel PKC inhibitor Gö6983 lowered the ionomycin-induced gain in cell surface GLUT4myc. Ionomycin stimulated GLUT4myc exocytosis and inhibited its endocytosis in live cells. siRNA-mediated knockdown of CaMKIIδ or AMPKα1/α2 partly reversed ionomycin-induced GLUT4myc exocytosis but did not prevent its reduced endocytosis. Compared with Gö6976, Gö6983 markedly reversed the slowing of GLUT4myc endocytosis triggered by ionomycin. In summary, rapid Ca(2+) influx into muscle cells accelerates GLUT4myc exocytosis while slowing GLUT4myc endocytosis. CaMKIIδ and AMPK stimulate GLUT4myc exocytosis, whereas novel PKCs reduce endocytosis. These results identify how Ca(2+)-activated signals selectively regulate GLUT4 exocytosis and endocytosis in muscle cells.

[Erythrokeratodermia variabilis]. / Érythrokératodermie variable.

BACKGROUND: Erythrokeratodermia variabilis (EKV) is a rare genodermatosis associated with keratinisation disorders. Mutations are found in genes encoding connexin 31 and 30.3 mapped to chromosome 1 p34-35. We report two cases of EKV, one of which presented dramatic improvement with oral retinoids. PATIENTS AND METHODS: A 15-month-old boy was referred to us with reddish-brown hyperkeratotic and well-demarcated plaques on the extremities, axillary space and face. The lesions started when he was 6months of age. Cutaneous histopathology showed acanthosis and papillomatosis associated with orthokeratotic hyperkeratosis. Anatomoclinical comparison confirmed the diagnosis of EKV. A second child aged 10years was referred to us with fixed, well-demarcated hyperkeratotic plaques associated with transient red patches. The lesions began when she was 1month old. Anatomoclinical comparison confirmed the diagnosis of EKV and the patient showed dramatic improvement after 2weeks on acitretin. DISCUSSION: EKV is characterized by the association of fixed well-demarcated plaques and transient erythematous patches. Although cutaneous histopathology is not specific, a typical physical examination and a compatible cutaneous histopathology can aid the diagnosis. Oral retinoids are often very rapidly effective.

Regulation of glucose transporter 4 traffic by energy deprivation from mitochondrial compromise.

Skeletal muscle is the major store and consumer of fatty acids and glucose. Glucose enters muscle through glucose transporter 4 (GLUT4). Upon insufficient oxygen availability or energy compromise, aerobic metabolism of glucose and fatty aids cannot proceed, and muscle cells rely on anaerobic metabolism of glucose to restore cellular energy status. An increase in glucose uptake into muscle is a key response to stimuli requiring rapid energy supply. This chapter analyses the mechanisms of the adaptive regulation of glucose transport that rescue muscle cells from mitochondrial uncoupling. Under these conditions, the initial drop in ATP recovers rapidly, through a compensatory increase in glucose uptake. This adaptive response involves AMPK activation by the initial ATP drop, which elevates cell surface GLUT4 and glucose uptake. The gain in surface GLUT4 involves different signals and routes of intracellular traffic compared with those engaged by insulin. The hormone increases GLUT4 exocytosis through phosphatidylinositol 3-kinase and Akt, whereas energy stress retards GLUT4 endocytosis through AMPK and calcium inputs. Given that energy stress is a component of muscle contraction, and that contraction activates AMPK and raises cytosolic calcium, we hypothesize that the increase in glucose uptake during contraction may also involve a reduction in GLUT4 endocytosis.

Reduction of insulin-stimulated glucose uptake in L6 myotubes by the protein kinase inhibitor SB203580 is independent of p38MAPK activity.

Insulin increases glucose uptake through translocation of the glucose transporter GLUT4 to the plasma membrane. We previously showed that insulin activates p38MAPK, and inhibitors of p38MAPKalpha and p38MAPKbeta (e.g. SB203580) reduce insulin-stimulated glucose uptake without affecting GLUT4 translocation. This observation suggested that insulin may increase GLUT4 activity via p38alpha and/or p38beta. Here we further explore the possible participation of p38MAPK through a combination of molecular strategies. SB203580 reduced insulin stimulation of glucose uptake in L6 myotubes overexpressing an SB203580-resistant p38alpha (drug-resistant p38alpha) but barely affected phosphorylation of the p38 substrate MAPK-activated protein kinase-2. Expression of dominant-negative p38alpha or p38beta reduced p38MAPK phosphorylation by 70% but had no effect on insulin-stimulated glucose uptake. Gene silencing via isoform-specific small interfering RNAs reduced expression of p38alpha or p38beta by 60-70% without diminishing insulin-stimulated glucose uptake. SB203580 reduced photoaffinity labeling of GLUT4 by bio-LC-ATB-BMPA only in the insulin-stimulated state. Unless low levels of p38MAPK suffice to regulate glucose uptake, these results suggest that the inhibition of insulin-stimulated glucose transport by SB203580 is likely not mediated by p38MAPK. Instead, changes experienced by insulin-stimulated GLUT4 make it susceptible to inhibition by SB203580.

Troglitazone causes acute mitochondrial membrane depolarisation and an AMPK-mediated increase in glucose phosphorylation in muscle cells.

AIMS/HYPOTHESIS: Troglitazone was the first thiazolidinedione (TZD) approved for clinical use, exerting hypoglycaemic effects related to its action as a ligand of the peroxisome proliferator-activated receptor gamma receptor in adipocytes. However, emerging evidence suggests that mitochondrial function may be affected by troglitazone, and that skeletal muscle cells acutely respond to troglitazone by enhancing glucose uptake. The aim of the present study was to determine the cellular mechanisms by which troglitazone acutely stimulates glucose utilisation in skeletal muscle cells. METHODS: L6 cells overexpressing GLUT4myc were incubated with troglitazone. Glucose uptake, transport and phosphorylation as well as AMP-activated protein kinase (AMPK) signalling and insulin signalling were examined. Changes in mitochondrial membrane potential were measured using the J-aggregate-forming dye JC-1. AMPK signalling was interfered with using AMPK alpha1/alpha2 siRNA. RESULTS: Troglitazone acutely (in 10 min) reduced the mitochondrial membrane potential in L6GLUT4myc myotubes and robustly stimulated AMPK activity. Following 30 min of incubation with troglitazone or insulin, 2-deoxyglucose uptake was stimulated 1.5- and 2.1-fold respectively, and in cells treated with troglitazone, a 1.8-fold increase in the 2-deoxyglucose-6-phosphate:2-deoxyglucose ratio was observed. Moreover, contrary to insulin, troglitazone did not significantly stimulate 3-O-methylglucose uptake. Unlike insulin, troglitazone did not increase surface GLUT4myc content and did not increase IRS1-associated phosphatidylinositol 3-kinase activity or Akt phosphorylation on T308 and S473. Interestingly, interfering with troglitazone-induced activation of AMPK by decreasing the expression of the enzyme using siRNA inhibited the stimulation of 2-deoxyglucose uptake by the TZD. CONCLUSIONS/INTERPRETATION: We propose that troglitazone acutely increases glucose flux in muscle via an AMPK-mediated increase in glucose phosphorylation.

Indinavir uncovers different contributions of GLUT4 and GLUT1 towards glucose uptake in muscle and fat cells and tissues.

AIMS/HYPOTHESIS: Insulin-dependent glucose influx in skeletal muscle and adipocytes is believed to rely largely on GLUT4, but this has not been confirmed directly. We assessed the relative functional contribution of GLUT4 in experimental models of skeletal muscle and adipocytes using the HIV-1 protease inhibitor indinavir. METHODS: Indinavir (up to 100 micro mol/l) was added to the glucose transport solution after insulin stimulation of wild-type L6 muscle cells, L6 cells over-expressing either GLUT4myc or GLUT1myc, 3T3-L1 adipocytes, isolated mouse brown or white adipocytes, and isolated mouse muscle preparations. RESULTS: 100 micro mol/l indinavir inhibited 80% of both basal and insulin-stimulated 2-deoxyglucose uptake in L6GLUT4myc myotubes and myoblasts, but only 25% in L6GLUT1myc cells. Cell-surface density of glucose transporters was not affected. In isolated soleus and extensor digitorum longus muscles, primary white and brown adipocytes, insulin-stimulated glucose uptake was inhibited 70 to 80% by indinavir. The effect of indinavir on glucose uptake was variable in 3T3-L1 adipocytes, averaging 45% and 67% inhibition of basal and maximally insulin-stimulated glucose uptake, respectively. In this cell, fractional inhibition of glucose uptake by indinavir correlated positively with the fold-stimulation of glucose uptake by insulin, and was higher with sub-maximal insulin concentrations. The latter finding coincided with an increase only in GLUT4, but not GLUT1, in plasma membrane lawns. CONCLUSION/INTERPRETATION: Indinavir is a useful tool to assess different functional contributions of GLUT4 to glucose uptake in common models of skeletal muscle and adipocytes.

A single amino acid substitution in herpes simplex virus type 1 VP16 inhibits binding to the virion host shutoff protein and is incompatible with virus growth.

In addition to its well-established role in the activation of herpes simplex virus immediate-early gene transcription, VP16 interacts with and downregulates the function of the virion host shutoff protein (vhs), thereby attenuating vhs-mediated destruction of viral mRNAs and translational arrest at late times of infection. We have carried out two-hybrid analysis in vivo and protein-protein interaction assays in vitro to identify determinants in VP16 necessary for interaction with vhs. The minimal amino-terminal subfragment of VP16 capable of binding to vhs encompassed residues 1 to 345. Alteration of a single leucine at position 344 to alanine (L344A) in the context of the amino-terminal fragment of VP16 containing residues 1 to 404 was sufficient to abolish interaction with vhs in vitro and in vivo. Leu344 could be replaced with hydrophobic amino acids (Ile, Phe, Met, or Val) but not by Asn, Lys, or Pro, indicating that hydrophobicity is an important property of binding to vhs. VP16 harboring a loss-of-function mutation at L344 was not compromised in its ability to interact with host cell factor (HCF-1) or to activate transcription of viral immediate-early genes in transient-transfection assays. Virus complementation assays using the VP16-null virus 8MA and the VP16/vhs double-mutant virus 8MAdeltaSma showed that VP16(L344A) was able to complement the growth of 8MAdeltaSma but not 8MA. Thus, a single point mutation in VP16 uncouples binding to vhs from other functions of VP16 required for virus growth and indicates that direct physical association between VP16 and vhs is necessary to sustain a productive infection.