Treating depression with botulinum toxin: a pooled analysis of randomized controlled trials.

INTRODUCTION: Botulinum toxin A (BTA) injection into the glabellar region is currently being studied as a treatment for major depressive disorder (MDD). Here we explore efficacy data of this novel approach in a pooled analysis. METHODS: A literature search revealed 3 RCTs on this topic. Individual patient data and clinical end points shared by these 3 trials were pooled and analyzed as one study (n=134) using multiple regression models with random effects. RESULTS: In the pooled sample, the BTA (n=59) and the placebo group (n=75) did not differ in the baseline variables. Efficacy outcomes revealed BTA superiority over placebo: Improvement in the Hamilton Depression Rating Scale or Montgomery-Asberg Depression Rating Scale 6 weeks after baseline was 45.7% for BTA vs. 14.6% for placebo (p<0.0001), corresponding to a BTA response rate of 54.2% (vs. 10.7%) and a BTA remission rate of 30.5% (vs. 6.7%). DISCUSSION: Equalling the status of a meta-analysis, this study increases evidence that a single treatment of BTA into the glabellar region can reduce symptoms of MDD. Further studies are needed to better understand how BTA exerts its mood-lifting effect.

Botulinum toxin therapy of bipolar depression: A case series.

We and others have recently found that botulinum toxin injected into the brow muscles has significant antidepressant properties as compared to placebo in randomized controlled trials in patients with major depressive disorder. However, data for the treatment of bipolar depression with botulinum toxin is lacking. We report here six patients with bipolar disorder experiencing moderate to severe depressive episodes who were treated on a compassionate basis with botulinum toxin given their persistent depressive symptoms and adverse side effects from medications. Four of six patients with bipolar depression experienced a remission following treatment with botulinum toxin, and the other two patients experienced a reduction of depressive symptoms. When the effect of botulinum toxin on the frown muscles began to wear off, depressive symptoms returned and retreatment with botulinum toxin provided successful relief of depressive symptoms again.

TGF-alpha is widely expressed in differentiated as well as hyperproliferative skin epithelium.

Transforming growth factor-alpha (TGF-alpha) is a potent mitogen for epithelial cells that is expressed at low levels in normal epidermis and overexpressed in psoriasis. Epidermal growth factor (EGF) has been shown to inhibit hair growth but stimulate the growth of sebaceous and sweat glands, suggesting a potential role for a member of the EGF/TGF-alpha family in the normal development and function of skin appendages as well as epidermis. The present work demonstrates TGF-alpha protein in eccrine ducts, and eccrine, sebaceous, and apocrine glands. The proliferative dermal hair bulb does not express TGF-alpha in contrast to the differentiated outer root sheath hair follicle epithelia. In addition, hyperproliferative skin diseases including bullous congenital ichthyosiform erythroderma, squamous cell carcinoma, and psoriasis show increased TGF-alpha expression. Thus, TGF-alpha may play a role in the morphogenesis and function of normal skin appendages and its overexpression is common in benign and malignant hyperproliferative skin diseases.

Retroviral expression of transforming growth factor-alpha does not transform fibroblasts or keratinocytes.

Transforming growth factor alpha (TGF alpha) is a peptide so named because it helps to impart anchorage-independent growth to normal rat kidney (NRK) cells in vitro and is secreted by many rodent and human tumor cells. To directly investigate the transforming properties of this factor, we constructed a replication-defective murine retrovirus that expresses the human sequence coding for TGF alpha. Infection of NIH/3T3 cells with the TGF alpha retrovirus led to the integration of a transcriptionally active provirus and overexpression of biologically active TGF alpha, but failed to induce morphologic transformation. Similarly, the TGF alpha retrovirus failed to induce morphologic transformation of five other types of rodent fibroblasts. We also investigated the effect of TGF alpha expression on the growth of BALB/MK mouse keratinocytes, which require epidermal growth factor (EGF) for proliferation. We show that exogenously added TGF alpha is an extremely potent mitogen for BALB/MK cells. However, retroviral expression of TGF alpha in BALB/MK cells failed to relieve dependence on exogenously added EGF (or TGF alpha) for cell growth. These results suggest that overexpression of TGF alpha does not, by itself, transform rodent fibroblasts or keratinocytes.

Transforming growth factor alpha expression helps to distinguish keratoacanthomas from squamous cell carcinomas.

Keratoacanthomas may be difficult to distinguish histologically from squamous cell carcinomas. We studied 20 keratocanthomas and 22 squamous cell carcinomas immunohistochemically using an antibody directed against transforming growth factor alpha to determine if the pattern of transforming growth factor alpha expression would provide a useful method of differentiating these tumors. Ninety percent of the keratoacanthomas demonstrated a diffuse pattern within tumor lobules in which all but the most peripheral rim of cells were stained. A similar localization of transforming growth factor alpha was not identified in squamous cell carcinomas. In addition, 40% of the squamous cell carcinomas but none of the keratoacanthomas showed focal transforming growth factor alpha immunostaining. Our results suggest that transforming growth factor alpha expression may be a marker of epithelial differentiation and may help distinguish between these two tumors.

The stimulation of yeast mitochondrial protein synthesis by low molecular weight cytoplasmic factors: requirement for intact mitochondria and lack of effect by folate derivatives.

Protein synthesis by GTP-supplemented yeast mitochondria is stimulated by a fraction of molecular weight less than 2,000 isolated from yeast high-speed supernatant (S-150). The low molecular weight fraction works independently of the respiratory chain as the stimulation effect is not cyanide-sensitive. Stimulation of mitochondrial protein synthesis by cytoplasmic factors is dependent upon the method of mitochondrial isolation. The low molecular weight stimulatory factor(s) are not reduced folate derivatives which supply formyl groups required for initiation of mitochondrial protein synthesis.

Effect of temperature on protein synthesis and leucine transport by yeast mitochondria.

Protein synthesis in yeast mitochondria shows biphasic Arrhenius plots both in vivo and in vitro, with a twofold increase in the activation energy below the transition temperature suggesting a functional association between mitochondrial protein synthesis and the inner membrane. Analysis by gel electrophoresis of mitochondrial translation products labeled in vivo showed that the same proteins are synthesized and then inserted into the membrane above and below the transition temperature of the membrane. The rate of leucine uptake into mitochondria was decreased at least fivefold in the presence of chloramphenicol, suggesting that leucine is used mainly for protein synthesis. In the absence of chloramphenicol, the rate of leucine uptake was always slightly higher but comparable to the incorporation rate of leucine into protein at all temperatures studied, suggesting that the transport of leucine into mitochondria is not rate-limiting for protein synthesis. The ionophore valinomycin or the uncoupler carbonyl phenylhydrazone (CCCP) inhibited 75-80% of the leucine uptake in the presence of chloramphenicol. In addition, the omission of respiratory chain substrates and the ATP-regenerating system led to a 93% inhibition of uptake, suggesting that leucine uptake may occur by an active transport mechanism.

Partial purification of cytosolic proteins which control yeast mitochondrial protein synthesis.

Protein synthesis in isolated yeast mitochondria incubated in the presence of GTP is stimulated 2-fold by addition of dialyzed postpolysomal supernatant (S-150) at the start of the incubation. Incubation of the yeast S-150 with 5'-nucleotidase had no effect on the stimulatory activity suggesting that the increased protein synthesis does not result from guanine nucleotides. A partial purification of the protein factors which stimulate mitochondrial protein synthesis has been accomplished by chromatography on Sephacryl S-200. Stimulatory activity was eluted in two peaks, one in the 40,000 to 80,000 molecular weight range and a broad peak with a molecular weight of less than 10,000. Stimulation of mitochondrial protein synthesis by the low molecular weight activator fraction was proportional to the concentration of protein added and abolished by trypsin treatment suggesting that the low molecular weight activator is a protein(s). The rate of mitochondrial protein synthesis in the presence of activator, was linear for 40 min, while that in the presence of GTP was linear for only 20 min, suggesting that the activator and GTP stimulate protein synthesis by different mechanisms. Analysis of the products of the stimulated mitochondrial protein synthesis by gel electrophoresis revealed that the activator increased equally the labeling of all products. These results indicate that low molecular weight proteins present in the cytosol regulate mitochondrial protein synthesis.

Cellular localization of retinoic acid receptor-gamma expression in normal and neoplastic skin.

Retinoids profoundly affect the normal growth and differentiation of epithelial tissues. Retinoic acid receptor-gamma (RAR-gamma) is a member of a family of retinoid receptors, and has been shown to be expressed almost exclusively in skin. However, little is known about the cellular localization of this receptor in human skin. The authors studied the expression of RAR-gamma in normal skin and human skin tumors by Northern blot analysis and in situ hybridization. RAR-gamma mRNA was detected in normal skin as well as in cultures of neonatal keratinocytes. Using an oligonucleotide specific for the RAR-gamma cDNA isoform 1 (RAR-gamma 1), RAR-gamma 1 mRNA was localized to all layers of the epidermis, the outer root sheath of hair follicles, follicular hair bulbs, eccrine and sebaceous glands. Basal cell carcinoma constitutively expressed gamma-1 mRNA and one of seven squamous cell carcinomas showed loss of gamma-1 mRNA expression, relative to adjacent epithelium. By contrast, normal melanocytic nevi and tumor-associated lymphocytes expressed little or no RAR-gamma mRNA. These results suggest that RAR-gamma 1 may play an important role in the maintenance and differentiation of normal epidermis and skin appendages.

Localization of transforming growth factor-alpha in human appendageal tumors.

Transforming growth factor-alpha (TGF alpha) is a potent mitogen for epithelial cells that has been localized to normal human appendageal epithelia. To further understand the role of TGF alpha in human appendages, we examined TGF alpha expression immunohistochemically in 17 types of human appendageal tumors differentiating toward hair follicles, eccrine, apocrine, and sebaceous glands. In order of decreasing degrees of differentiation, tumors could be divided into hyperplasias, adenomas, benign epitheliomas, and primordial epitheliomas. Using an antibody that recognizes primarily the 6-kd and 13-kd forms of TGF alpha, TGF alpha immunostaining in 16 of 17 tumor types analyzed was found to follow a similar pattern, with expression in hyperplasias greater than adenomas greater than benign epitheliomas greater than primordial epitheliomas. Within a given tumor, TGF alpha expression also correlated well with the known differentiation state of the tumor cell types. The results suggest that TGF alpha expression is directly correlated with the differentiation state of hair follicle, eccrine, apocrine, and sebaceous tumors in human skin, and raises the possibility that TGF alpha may play a role in the differentiation of appendageal epithelia.

Role of genes for normal growth factors in human malignancy.

The human homologue of the v-sis oncogene encodes one chain of human platelet-derived growth factor (PDGF-2). Previous studies have shown that expression of the coding sequence for this growth factor induces malignant transformation of NIH/3T3 cells. We demonstrate the detection of sis/PDGF-2 products indistinguishable from functional PDGF-2 homodimers in human tumor cells. These findings support the concept that expression of the sis/PDGF-2 product in human cells responsive to its proliferative actions can be an important step in the processes leading to malignancy. Unlike sis/PDGF-2, which remains tightly cell-associated, another growth factor, termed transforming growth factor alpha (TGF alpha), is actively secreted. Expression vectors for the TGF alpha coding sequence failed to induce primary transformed foci upon transfection of NIH/3T3 cells despite high levels of TGF alpha synthesized by these cells. Moreover, transfectants selected for secretion of high levels of TGF alpha failed to form colonies on contact inhibited NIH/3T3 monolayers or to induce tumors upon inoculation of nude mice. Thus, the ability of a growth factor to induce autonomous in vitro or in vivo growth is dependent upon more than expression of cognate receptors by the cell in which it is synthesized.

The human transforming growth factor type alpha coding sequence is not a direct-acting oncogene when overexpressed in NIH 3T3 cells.

A peptide secreted by some tumor cells in vitro imparts anchorage-independent growth to normal rat kidney (NRK) cells and has been termed transforming growth factor type alpha (TGF-alpha). To directly investigate the transforming properties of this factor, the human sequence coding for TGF-alpha was placed under the control of either a metallothionein promoter or a retroviral long terminal repeat. These constructs failed to induce morphological transformation upon transfection of NIH 3T3 cells, whereas viral oncogenes encoding a truncated form of its cognate receptor, the EGF receptor, or another growth factor, sis/platelet-derived growth factor 2, efficiently induced transformed foci. When NIH 3T3 clonal sublines were selected by transfection of TGF-alpha expression vectors in the presence of a dominant selectable marker, they were shown to secrete large amounts of TGF-alpha into the medium, to have downregulated EGF receptors, and to be inhibited in growth by TGF-alpha monoclonal antibody. These results indicated that secreted TGF-alpha interacts with its receptor at a cell surface location. Single cell-derived TGF-alpha-expressing sublines grew to high saturation density in culture. However, when plated as single cells on contact-inhibited monolayers of NIH 3T3 cells, they failed to form colonies, whereas v-sis- and v-erbB-transfected cells formed transformed colonies under the same conditions. Moreover, TGF-alpha-expressing sublines were not tumorigenic in nude mice. These and other results imply that TGF-alpha exerts a growth-promoting effect on the entire NIH 3T3 cell population after secretion into the medium but little, if any, effect on the individual cell synthesizing this factor. It is concluded that the normal coding sequence for TGF-alpha is not a direct-acting oncogene when overexpressed in NIH 3T3 cells.

TGF alpha stimulates growth of skin papillomas by autocrine and paracrine mechanisms but does not cause neoplastic progression.

To investigate the role of transforming growth factor alpha (TGF alpha) in tumor development, we introduced the human TGF alpha (hTGF alpha) cDNA into cultured primary mouse epidermal cells or papilloma cells using a replication-defective retroviral vector and analyzed skin grafts constructed with such cells. Expression of the exogenous gene was confirmed by detection of hTGF alpha mRNA by northern RNA blot analysis, and the secreted hTGF alpha was measured by ELISA of culture supernatants. Tumor cells expressing hTGF alpha produced benign tumors (papillomas), which were 1.5- to 2-fold larger than tumors of parental cells when tested as skin grafts on nude mice. Grafts of normal cells that expressed hTGF alpha produced normal skin. When mixtures of parental tumor cells and normal mouse keratinocytes were grafted to nude mice, papilloma formation was suppressed and tumors that did form were small. Grafts of hTGF alpha-producing papilloma cells combined with either normal epidermal cells or hTGF alpha-producing epidermal cells yielded large tumors. Mixed grafts containing keratinocytes expressing hTGF alpha and parental papilloma cells also produced large tumors. While the tumor size was substantially increased by hTGF alpha expression, the tumors that developed in all groups were histologically benign and reached a stable size 4-5 wk after grafting. These results indicate that expression of hTGF alpha by either tumor cells (autocrine) or adjoining normal cells (paracrine) can stimulate tumor growth, particularly when tumor growth is suppressed by normal tissue. However, expression of this growth factor did not appear to influence tumor progression directly.